Therapeutic agent for FGFR inhibitor-resistant cancer

ABSTRACT

The problem to be solved by the present invention is to provide an anticancer agent for treating tumors resistant to other antitumor agents that inhibit FGFR, and a method for treating such tumors. The present invention provides an antitumor agent for administration to a tumor patient resistant to an FGFR inhibitor, the antitumor agent comprising a 3,5-disubstituted benzene alkynyl compound represented by Formula (I) below or a salt thereof. The present invention also provides a therapeutic method using the anticancer agent.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a continuation of U.S. Ser. No. 14/905,047, filedJan. 14, 2016, which is a 35 U.S.C. 371 National Phase Entry Applicationfrom PCT/JP2014/069105, filed Jul. 17, 2014, which claims priority basedon Japanese Patent Application No. 2013-149958 filed on Jul. 18, 2013,the entire contents of which are incorporated herein by reference.

TECHNICAL FIELD

The present invention relates to an antitumor agent comprising an FGFRinhibitor as an active ingredient.

BACKGROUND ART

Fibroblast growth factors (FGFs) are involved in the regulation ofvarious physiological processes, such as cell proliferation, chemotaxis,and differentiation. The physiological activity of the FGFs is mediatedby fibroblast growth factor receptors (FGFRs), which are specific cellsurface receptors. FGFRs belong to a receptor protein tyrosine kinasefamily, and comprise an extracellular ligand-binding domain, a singletransmembrane domain, and an intracellular tyrosine kinase domain. Fourtypes of FGFRs (FGFR1, FGFR2, FGFR3, and FGFR4) have been heretoforeidentified. FGFRs bind to FGFs to form dimers, and are activated byphosphorylation. Activation of the receptors induces dimerization andactivation of specific downstream signal transduction molecules, therebydeveloping physiological functions.

Many reports have been made about the relationship between aberrantFGF/FGFR signaling and various human cancers (e.g., NPL 1, NPL 2, andNPL 3). Aberrant activation of FGF/FGFR signaling in human cancer isconsidered to be attributable to overexpression of FGFRs and/or geneamplification, gene mutation, chromosomal translocation, or an autocrineor paracrine mechanism by overproduction of FGFs (ligands). Moreover,such aberrant signaling is considered to be partly responsible fortherapeutic resistance to existing chemotherapeutic anticancer drugs orother receptor tyrosine kinase inhibitors in human cancer (NPL 4).Furthermore, the aberrant signaling is known to be associated withvarious diseases caused by abnormal angiogenic processes, such as solidtumor, rheumatoid arthritis, psoriasis, retinopathy, and age-relatedmacular degeneration (NPL 5).

Accordingly, therapies targeted for FGF/FGFR signaling not only have adirect antitumor effect on tumor cells that are highly dependent onFGF/FGFR signaling, but also exhibit an inhibitory effect on tumorangiogenesis induced by FGF/FGFR signaling; thus, such therapies areexpected to be promising targeted therapies having sufficient antitumoreffects. In addition, such therapies are expected to provide drug effectenhancers for existing chemotherapeutic anticancer drugs or otherreceptor tyrosine kinase inhibitors, or effective therapeutic remediesfor cancer types that are resistant or unresponsive to these drugs.

PTL 1 discloses a wide range of fused bicyclic compounds having mTORinhibitory activity; however, the specifically disclosed compounds areall imidazopyrazine compounds, and FGFR inhibitory activity is nowherementioned in PTL 1. PTL 2 discloses BTK inhibitor compounds having acharacteristic substituent at the 3-position of the pyrazolopyrimidinering, but is silent about FGFR inhibitory activity. PTL 3 disclosesHSP90 inhibitor compounds having a characteristic substituent at the5-position of the pyrrolopyrimidine ring, but is silent about FGFRinhibitory activity.

Further, in the treatment of a tumor using an FGFR inhibitor, the tumorbecomes resistant to the inhibitor due to FGFR mutations, which mayadversely affect the prognosis of the tumor patient (PTL 4).

CITATION LIST Patent Literature

-   PTL 1: WO 2007/087395-   PTL 2: WO 2008/121742-   PTL 3: WO 2010/043865-   PTL 4: WO 2011/115937

Non-Patent Literature

-   NPL 1: J. Clin. Oncol. 24, 3664-3671 (2006)-   NPL 2: Mol. Cancer Res. 3, 655-667 (2005)-   NPL 3: Cancer Res. 70, 2085-2094 (2010)-   NPL 4: Clin. Cancer Res. 17, 6130-6139 (2011)-   NPL 5: Nat. Med. 1, 27-31 (1995)

SUMMARY OF INVENTION Technical Problem

The problem to be solved by the present invention is to provide ananticancer agent for treating tumors resistant to other antitumor agentsthat inhibit FGFR, and a method for treating such tumors.

Solution to Problem

The present inventors further conducted various studies on methods fortreating tumors resistant to conventional FGFR inhibitors, andconsequently found that a 3,5-disubstituted benzene alkynyl compound ora salt thereof exhibits antitumor effects, and is very effective as amethod for treating such tumors. The present invention is based on thisnovel finding.

Therefore, the present invention provides the following items:

Item 1. An antitumor agent for administration to a tumor patientresistant to an FGFR inhibitor, the antitumor agent comprising a3,5-disubstituted benzene alkynyl compound represented by Formula (I):

-   -   or a salt thereof,    -   wherein R₁ is the same or different, and each represents C₁-C₆        alkyl;    -   X₁ and X₂ independently represent N or CH;    -   Y is a group represented by Formula (A):

-   -   -   wherein the divalent moiety represented by

-   -   -   is a nitrogen-containing C₃-C₁₀ heterocycloalkylene group, a            group represented by Formula (B):

-   -   -   wherein the divalent moiety represented by

-   -   -   is a C₃-C₁₀ cycloalkylene group, or

    -   a group represented by Formula (C):

-   -   -   wherein the divalent moiety represented by

-   -   -   is a C₆-C₁₂ arylene group;

    -   R₂ is hydrogen, C₂-C₆ alkynyl, —C(═O)OR_(x),        —C(═O)N(R_(x))(R_(y)), hydroxy-C₁-C₆ alkyl, di(C₁-C₆        alkyl)amino-C₁-C₆ alkyl, or C₂-C₉ heteroaryl optionally having        R₃; and

    -   R₃ is C₁-C₆ alkyl or di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl;

    -   Z is —C(R₄)═C(R₅) (R₆) or —C≡C—R₇;

    -   R₄, R₅, and R₆ are the same or different, and each represents        hydrogen, halogen, C₁-C₆ alkyl optionally having R₈, or a group        represented by Formula (D):

-   -   -   wherein the monovalent moiety represented by

-   -   -   is a nitrogen-containing C₃-C₁₀ heterocycloalkyl group,

    -   R₇ is hydrogen, C₁-C₆ alkyl, or hydroxy-C₁-C₆ alkyl;

    -   R₈ is —OR_(x) or —N(R_(x)) (R_(y));

    -   R₉ is C₁-C₆ alkyl, halogen, or —OR_(x);

    -   R_(x) and R_(y) are the same or different, and each represents        hydrogen, C₁-C₆ alkyl, C₃-C₁₀ cycloalkyl, di(C₁-C₆        alkyl)amino-C₁-C₆ alkyl, or C₁-C₆ alkoxy-C₁-C₆ alkyl;

    -   l is an integer of 0 to 3;

    -   m is an integer of 1 to 3; and n is an integer of 0 to 2.

Item 2. A method for treating a tumor patient resistant to an FGFRinhibitor, the method comprising administering an antitumor agentcomprising a 3,5-disubstituted benzene alkynyl compound represented byFormula (I) above, wherein R₁, X₁, X₂, Y, and Z are as defined above, ora salt thereof.

Item 3. A 3,5-disubstituted benzene alkynyl compound represented byFormula (I) above, wherein R₁, X₁, X₂, Y, and Z are as defined above, ora salt thereof for treatment of a tumor patient resistant to an FGFRinhibitor.

Item 4. Use of a 3,5-disubstituted benzene alkynyl compound representedby Formula (I) above, wherein R₁, X₁, X₂, Y, and Z are as defined above,or a salt thereof for producing an antitumor agent for administration toa tumor patient resistant to an FGFR inhibitor.

Item 5. The antitumor agent according to item 1, the method according toitem 2, the compound or a salt thereof according to item 3, or the useaccording to item 4, wherein in Formula (I), (1) when X₂ is N, X₁ is Nor CH, and (2) when X₂ is CH, X₁ is CH.

Item 6. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 5, wherein in Formula (I), l is 0 or1.

Item 7. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 6, wherein in Formula (I), (1) when Yis a group represented by Formula (A), the group represented by

is azetidinylene, pyrrolidinylene, piperidinylene, piperazinylene, ormorpholinylene, (2) when Y is a group represented by Formula (B), thegroup represented by

is cyclopropylene or cyclobutylene, and (3) when Y is a grouprepresented by Formula (C), the group represented by

is phenylene.

Item 8. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 7, wherein in Formula (I), (1) when Yis a group represented by Formula (A), Z is —C(R₄)═C(R₅)(R₆) or —C≡C—R₇,and (2) when Y is a group represented by Formula (B) or (C), Z is—C(R₄)═C(R₅)(R₆).

Item 9. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 8, wherein in Formula (I), R₁ ismethyl or ethyl.

Item 10. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 9, wherein in Formula (I), when X₂ isN, X₁ is N or CH, and (2) when X₂ is CH, X₁ is CH; l is 0 or 1; and (1)when Y is a group represented by Formula (A), the group represented by

is azetidinylene, pyrrolidinylene, piperidinylene, piperazinylene, ormorpholinylene, (2) when Y is a group represented by Formula (B), thegroup represented by

is cyclopropylene or cyclobutylene, and (3) when Y is a grouprepresented by Formula (C), the group represented by

is phenylene.

Item 11. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 8, wherein the 3,5-disubstitutedbenzene alkynyl compound is selected from the following group ofcompounds:

-   (1)    (S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one,-   (2)    (S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-yn-1-one,-   (3)    (S)-1-(3-(4-amino-3-((3,5-diethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one,-   (4)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)prop-2-en-1-one,-   (5)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-hydroxybut-2-yn-1-one,-   (6)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(dimethylamino)but-2-en-1-one,-   (7)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(cyclopropylamino)but-2-en-1-one,-   (8)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(isopropylamino)but-2-en-1-one,-   (9)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(ethyl(methyl)amino)but-2-en-1-one,-   (10)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(cyclobutylamino)but-2-en-1-one,-   (11)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(diethylamino)but-2-en-1-one,-   (12)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(tert-butylamino)but-2-en-1-one,-   (13)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(isopropyl(methyl)amino)but-2-en-1-one,-   (14)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(piperidin-1-yl)but-2-en-1-one,-   (15)    (S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(3-fluoropyrrolidin-1-yl)but-2-en-1-one,-   (16)    (R)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(3-fluoropyrrolidin-1-yl)but-2-en-1-one,-   (17)    1-((2S,4S)-4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-(hydroxymethyl)pyrrolidin-1-yl)prop-2-en-1-one,-   (18)    1-(2S,4S)-4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynylpyrrolidin-1-yl)prop-2-en-1-one,-   (19)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one,-   (20)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)prop-2-en-1-one,-   (21)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(pyrrolidin-1-yl)but-2-en-1-one,-   (22)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(4-hydroxypiperidin-1-yl)but-2-en-1-one,-   (23)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)but-2-yn-1-one,-   (24)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-hydroxy-4-methylpent-2-yn-1-one,-   (25)    1-((S)-3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-((S)-3-fluoropyrrolidin-1-yl)but-2-en-1-one,-   (26)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(piperidin-1-yl)but-2-en-1-one,-   (27)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)prop-2-en-1-one,-   (28)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(dimethylamino)but-2-en-1-one,-   (29)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(pyrrolidin-1-yl)but-2-en-1-one,-   (30)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(azetidin-1-yl)but-2-en-1-one,-   (31)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(ethyl(methyl)amino)but-2-en-1-one,-   (32)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(isopropylamino)but-2-en-1-one,-   (33)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(diethylamino)but-2-en-1-one,-   (34)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-((2-methoxyethyl)(methyl)amino)but-2-en-1-one,-   (35)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(4-hydroxypiperidin-1-yl)but-2-en-1-one,-   (36)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(3-hydroxypyrrolidin-1-yl)but-2-en-1-one,-   (37)    (R)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(3-hydroxypyrrolidin-1-yl)but-2-en-1-one,-   (38)    (2S,4S)-methyl-1-acryloyl-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate,-   (39)    1-((2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(1,3,4-oxadiazol-2-yl)pyrrolidin-1-yl)prop-2-en-1-one,    and-   (40)    (S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one.

Item 12. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 11, wherein the 3,5-disubstitutedbenzene alkynyl compound or a salt thereof is(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-oneor a salt thereof.

Item 13. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 12, wherein the tumor patient isresistant to at least one of ponatinib, regorafenib, intedanib,dovitinib lactate, lenvatinib mesylate, cediranib, oratinib, brivanibalaninate, AZD4547, NVP-BGJ398, sulfatinib, ARQ-087, S-49076, IMCA1,PRO001, and R3Mab.

Item 14. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 13, wherein the tumor patient has amutation at at least one of N550, V565, E566, and K660 of FGFR2.

Item 15. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 13, wherein the tumor patient has amutation at N550 of FGFR2.

Item 16. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 13, wherein the tumor patient has amutation at V565 of FGFR2.

Item 17. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 13, wherein the tumor patient has amutation at E566 of FGFR2.

Item 18. The antitumor agent, method, compound or a salt thereof, or useaccording to any one of items 1 to 13, wherein the tumor patient has amutation at K660 of FGFR2.

Advantageous Effects of Invention

It was revealed that the compound represented by Formula (I) above or asalt thereof, which is the active ingredient of the present invention,unexpectedly exhibited antitumor effects on tumors resistant toconventional FGFR inhibitors. Therefore, the compound represented byFormula (I) or a salt thereof is very effective for, among varioustumors, tumors resistant to conventional FGFR inhibitors.

BRIEF DESCRIPTION OF DRAWING

FIG. 1 shows the amino acid sequence (SEQ ID NO: 1) encoded by GenBankaccession number NP_075259.

DESCRIPTION OF EMBODIMENTS

The present invention provides an antitumor agent comprising a3,5-disubstituted benzene alkynyl compound represented by Formula (I)above or a salt thereof for administration to tumor patients resistantto FGFR inhibitors.

The tumor resistant to an FGFR inhibitor to be prevented and/or treatedin the present invention refers to a tumor that responded to aconventionally known FGFR inhibitor, but has become resistant to theantitumor effect of the inhibitor due to the continuous administrationthereof, and refers to a tumor that is originally resistant to aconventionally known FGFR inhibitor. Conventional FGFR inhibitors are,for example, ponatinib, regorafenib, intedanib, dovitinib lactate,lenvatinib mesylate, cediranib, oratinib, brivanib alaninate, AZD4547,NVP-BGJ398, sulfatinib, ARQ-087, S-49076, IMCA1, PRO001, R3Mab, and thelike; and typically AZD4547, NVP-BGJ398, and the like. Therefore, in thepresent invention, examples of tumors resistant to FGFR inhibitorsinclude tumors resistant to at least one of these FGFR inhibitors.

Examples of tumor patients resistant to FGFR inhibitors in the presentinvention include patients with a mutation at at least one of N550,V565, E566, and K660 of FGFR2. In particular, V565 corresponds to acharacteristic amino acid residue called a “gatekeeper” in the kinase.Patients with a mutation at the gatekeeper residue are also included. Inthis specification, the term “FGFR2” includes a protein having an aminoacid sequence (SEQ ID NO: 1; FIG. 1) encoded by GenBank accession numberNP_075259, a homologue, isoform, mutant, or derivative thereof, aprotein cut from the precursor protein, and the like. In the presentinvention, the position of the FGFR2 mutation denotes a positioncorresponding to the position in the amino acid sequence represented bySEQ ID NO: 1. The “mutation at N550” in FGFR2 indicates that asparaginein FGFR2 at a position corresponding to the 550th amino acid of SEQ IDNO: 1 is mutated. Examples of such FGFR2 mutations include N550H, V565I,E566G, K660M, and the like. Therefore, even if the mutation position ina certain FGFR2 is different from the mutation position of the aminoacid number encoded by NP_075259 due to the presence of splicingvariants, deletions, insertions, etc., the mutation at the positioncorresponding to the amino acid sequence encoded by NP_075259 is alsoincluded.

The compound represented by Formula (I) above, which is the activeingredient of the antitumor agent of the present invention, is a3,5-disubstituted benzene alkynyl compound containing a condensedheteroaryl group substituted for an α,β-unsaturated amide via a spacermoiety, and is not disclosed in any of the above prior art documents.

In the present specification, the term “C₁-C₆ alkyl” refers to astraight or branched alkyl group having 1 to 6 carbon atoms. Specificexamples thereof include methyl, ethyl, n-propyl, isopropyl, n-butyl,isobutyl, sec-butyl, tert-butyl, pentyl, hexyl, and the like. The C₁-C₆alkyl is preferably a straight or branched alkyl group having 1 to 4carbon atoms (a C₁-C₄ alkyl group), and more preferably methyl, ethyl,isopropyl, and tert-butyl.

In this specification, the term “C₃-C₁₀ cycloalkyl” refers to amonocyclic or polycyclic cycloalkyl group having 3 to 10 carbon atoms,and is preferably a monocyclic cycloalkyl group having 3 to 6 carbonatoms (a C₃-C₆ cycloalkyl group). Specific examples thereof includecyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, decalyl,and the like. Cyclopropyl and cyclobutyl are preferable. In thisspecification, the divalent moiety represented by

of the group represented by Formula (A)

(wherein R₂ and l are as defined above)is a C₃-C₁₀ divalent heterocycloalkylene group containing at least onenitrogen atom in the ring and further containing 0 to 2 same ordifferent heteroatoms selected from oxygen and sulfur atoms in the ring(a nitrogen-containing C₃-C₁₀ heterocycloalkylene group), and ispreferably a C₃-C₅ heterocycloalkylene group containing 1 to 3 nitrogenatoms in the ring and further containing 0 to 1 oxygen atom in the ring(a nitrogen-containing C₃-C₅ heterocycloalkylene group). Specificexamples thereof include azetidinylene, pyrrolidinylene, piperidinylene,piperazinylene, morpholinylene, octahydroquinolinylene,octahydroindolylene, and the like. Among them, azetidinylene,pyrrolidinylene, piperidinylene, piperazinylene, and morpholinylene arepreferable.

The group represented by Formula (A)

refers to a divalent nitrogen-containing C₃-C₁₀ heterocycloalkylenegroup represented by

wherein the nitrogen atom has one arm and the other arm is connected toa substituent (—(CH₂)₁—), and a substituent R₂ is present on the ring.

In this specification, the divalent moiety represented by

of the group represented by Formula (B)

(wherein R₂ and l are as defined above)refers to a monocyclic or polycyclic divalent cycloalkylene group having3 to 10 carbon atoms (a C₃-C₁₀ cycloalkylene group), and preferably amonocyclic divalent cycloalkylene group having 3 to 6 carbon atoms (aC₃-C₆ cycloalkylene group). Specific examples thereof includecyclopropylene, cyclobutylene, cyclopentylene, cyclohexylene,cycloheptylene, decalylene, and the like. Cyclopropylene and (1,2- or1,3-)cyclobutylene are preferable.

refers to a divalent C₃-C₁₀ cycloalkylene group represented by

wherein one arm is connected to an adjacent amino group (NH) and theother arm is connected to a substituent (—(CH₂)₁—), and a substituent R₂is present on the ring.

In the present specification, the divalent moiety represented by

of the group represented by Formula (C)

(wherein R₂ and l are as defined above)refers to a divalent arylene group having 6 to 12 carbon atoms (a C₆-C₁₂arylene group). Specific examples thereof include phenylene,naphthylene, biphenylene, and the like. Phenylene is preferable.

refers to a divalent C₆-C₁₂ arylene group represented by

wherein one arm is connected to an adjacent amino group (NH) and theother arm is connected to a substituent (—(CH₂)₁—), and a substituent R₂is present on the ring.

In this specification, the monovalent moiety represented by

of the group represented by Formula (D)

(wherein R₉, m, and n are as defined above)refers to a C₃-C₁₀ heterocycloalkyl group containing at least onenitrogen atom in the ring and further containing 0 to 2 same ordifferent heteroatoms selected from oxygen and sulfur atoms in the ring(a nitrogen-containing C₃-C₁₀ heterocycloalkyl group), and is preferablya C₃-C₅ heterocycloalkyl group containing 1 to 3 nitrogen atoms in thering and further containing 0 to 1 oxygen atom in the ring (anitrogen-containing C₃-C₅ heterocycloalkyl group). Specific examplesthereof include azetidinyl, pyrrolidinyl, piperidinyl, piperazinyl,morpholinyl, octahydroquinolinyl, octahydroindolyl, and the like.Azetidinyl, pyrrolidinyl, piperidinyl, piperazinyl, and morpholinyl arepreferable.

denotes a nitrogen-containing C₃-C₁₀ heterocycloalkyl group representedby

wherein the nitrogen atom is bound to a substituent (—CH₂)_(n)—), and nsubstituents (—(R₉)_(n)) are present on the ring.

In this specification, the “C₂-C₉ heteroaryl” refers to a monocyclic orbicyclic C₂-C₉ heteroaryl group containing 1 to 3 same or differentheteroatoms selected from nitrogen, oxygen, and sulfur atoms; and ispreferably a monocyclic C₂-C₅ heteroaryl group containing 1 to 3 same ordifferent heteroatoms selected from nitrogen, oxygen, and sulfur atoms(a C₂-C₅ heteroaryl group). Specific examples thereof include thienyl,furyl, pyrrolyl, triazolyl, imidazolyl, pyrazolyl, oxadiazolyl,isothiazolyl, isoxazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl,isobenzofuryl, indolizinyl, isoindolyl, indolyl, indazolyl, quinolyl,isoquinolyl, phthalazinyl, naphthyridinyl, and the like.1,3,4-Oxadiazolyl is preferable.

In this specification, the term “C₂-C₆ alkynyl” refers to a straight orbranched C₂-C₆ alkynyl group having at least one carbon-carbon triplebond. Specific examples thereof include ethynyl, 2-propynyl, 2-hexynyl,and the like. Ethynyl is preferable.

In this specification, the term “hydroxy-C₁-C₆ alkyl” refers to astraight or branched C₁-C₆ alkyl group having one hydroxy group.Specific examples thereof include hydroxymethyl, hydroxyethyl,hydroxypropyl, hydroxybutyl, hydroxypentyl, hydroxyhexyl, and the like.Among them, hydroxymethyl, 2-hydroxyethyl, 2-hydroxypropyl, and2-hydroxybutyl are preferable.

In this specification, the term “di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl”refers to a straight or branched C₁-C₆ alkyl group having an amino grouphaving two straight or branched C₁-C₆ alkyl groups. A straight orbranched C₁-C₄ alkyl group having an amino group having two straight orbranched C₁-C₄ alkyl groups (a di(C₁-C₄ alkyl)amino-C₁-C₄ alkyl group)is preferable. Specific examples thereof include dimethylaminomethyl,dimethylaminoethyl, dimethylaminopropyl, dimethylaminobutyl,dimethylaminopentyl, dimethylaminohexyl, diethylaminomethyl,diethylaminoethyl, diethylaminopropyl, diethylaminobutyl,diethylaminopentyl, diethylaminohexyl, dipropylaminomethyl,dibutylaminomethyl, dipentylaminomethyl, dihexylaminomethyl,ethyl(methyl)aminomethyl, and the like. Dimethylaminomethyl anddiethylaminomethyl are preferable.

In this specification, the term “C₁-C₆ alkoxy-C₁-C₆ alkyl” refers to astraight or branched C₁-C₆ alkyl group having a straight or branchedC₁-C₆ alkoxy group. It is preferably a straight or branched C₁-C₄ alkylgroup having a straight or branched C₁-C₄ alkoxy group (a C₁-C₄alkoxy-C₁-C₄ alkyl group). Specific examples of such groups includemethoxymethyl, methoxyethyl, methoxypropyl, methoxybutyl, methoxypentyl,methoxyhexyl, ethoxymethyl, ethoxyethyl, ethoxypropyl, ethoxybutyl,ethoxypentyl, ethoxyhexyl, propoxymethyl, butoxymethyl, pentyloxymethyl,hexyloxymethyl, and the like. Among them, 2-methoxyethyl is preferable.

In this specification, examples of the “halogen” include chlorine,bromine, fluorine, and iodine. Fluorine is preferable.

In Formula (I), the following combinations of X₁ and X₂ are preferable.(1) When X₂ is N, X₁ is N or CH. (2) When X₂ is CH, X₁ is CH.

In Formula (I), l is preferably 0 or 1.

In Formula (I), Y is preferably a group represented by Formula (A)

(wherein R₂ and l are as defined above) or a group represented byFormula (C)

(wherein R₂ and l are as defined above). More preferably, the divalentmoiety represented by

of a group represented by Formula (A) is pyrrolidinylene, azetidinylene,or piperidinylene, or the divalent moiety represented by

of a group represented by Formula (C) is phenylene.

In Formula (I), the following combinations of Y and Z are preferable.When Y is a group represented by Formula (A)

(wherein R₂ and l are as defined above), Z is —C(R₄)═C(R₅) (R₆) or—C≡C—R₇. When Y is a group represented by Formula (B) or (C):

(wherein R₂ and l are as defined above), Z is —C(R₄)═C(R₅) (R₆).

In Formula (I), R₁ is preferably C₁-C₄ alkyl, and more preferably methylor ethyl.

In Formula (I), R₂ is preferably hydrogen, C₂-C₆ alkynyl, —C(═O)OR_(x),hydroxy-C₁-C₄ alkyl, or C₂-C₉ heteroaryl optionally having R₃, and morepreferably ethynyl, methoxycarbonyl, hydroxymethyl, or 1,3,4-oxadiazolyloptionally having R₃.

In Formula (I), R₃ is preferably C₁-C₄ alkyl or di-(C₁-C₄alkyl)amino-C₁-C₄ alkyl, and more preferably methyl ordimethylaminomethyl.

In Formula (I), R₄ is preferably hydrogen or halogen, more preferablyhydrogen or fluorine, and even more preferably hydrogen.

In Formula (I), R₅ and R₆ are preferably hydrogen, C₁-C₄ alkyl groupoptionally having R₈, or a group represented by Formula (D)

(wherein R₉, m and n are as defined above), and more preferablyhydrogen, methyl having R₈, or a group represented by Formula (D)

(wherein R₉, m, and n are as defined above).

In Formula (I), m is preferably 1.

In Formula (I), R₉ is preferably C₁-C₄ alkyl, fluorine, or hydroxy, andmore preferably methyl, fluorine, or hydroxy.

In Formula (I), n is preferably 0 or 1.

In Formula (I), R₇ is preferably hydrogen, C₁-C₄ alkyl, or hydroxy-C₁-C₄alkyl, and more preferably hydrogen, hydroxymethyl, methyl, or2-hydroxy-2-methyl-ethyl.

In Formula (I), R₈ is preferably hydroxy or —N(R_(x)) (R_(y)). In thisformula, R_(x) and R_(y) are preferably hydrogen, C₁-C₄ alkyl, C₃-C₁₀cycloalkyl, or C₁-C₄ alkoxy-C₁-C₄ alkyl, and more preferably hydrogen,methyl, ethyl, tert-butyl, isopropyl, cyclopropyl, cyclobutyl, or2-methoxyethyl.

Preferable Compounds (I) are compounds represented by Formula (I)wherein R₁ is C₁-C₄ alkyl; X₁ and X₂ are independently N or CH; Y is agroup represented by Formula (A) or (C):

R₂ is hydrogen, C₂-C₆ alkynyl, —C(═O)OR_(x), hydroxy-C₁-C₄ alkyl, orC₂-C₉ heteroaryl optionally having R₃; R₃ is C₁-C₄ alkyl or di(C₁-C₄alkyl)amino-C₁-C₄ alkyl; Z is —C(R₄)═C(R₅) (R₆) or —C≡C—R₇; R₄ ishydrogen or halogen; R₅ and R₆ are the same or different, and eachrepresents hydrogen, C₁-C₄ alkyl optionally having R₈, or a grouprepresented by Formula (D)

R₇ is hydrogen, C₁-C₄ alkyl, or hydroxy-C₁-C₄ alkyl; R₈ is hydroxy or—N(R_(x))(R_(y)); R₉ is C₁-C₄ alkyl, fluorine, or hydroxy; R_(x) andR_(y) are the same or different, and each represents hydrogen, C₁-C₄alkyl, C₃-C₁₀ cycloalkyl, or C₁-C₄ alkoxy-C₁-C₄ alkyl; and l is 0 or 1,m is 1, and n is 0 or 1.

More preferable Compounds (I) are compounds represented by Formula (I)wherein R₁ is C₁-C₄ alkyl; X₁ and X₂ are such that (1) when X₂ is N, X₁is N or CH, and (2) when X₂ is CH, X₁ is CH; in Y, the divalent moietyrepresented by

of the group represented by Formula (A)

is pyrrolidinylene, azetidinylene, or piperidinylene, or the divalentmoiety represented by

of the group represented by Formula (C)

is phenylene;(a) when Y is a group represented by Formula (A)

(wherein R₂ is hydrogen, ethynyl, methoxycarbonyl, hydroxymethyl, or1,3,4-oxadiazolyl optionally having R₃; R₃ is C₁-C₄ alkyl; and 1 is 0 or1), Z is —C(R₄)═C(R₅)(R₆) or —C≡C—R₇, (b) when Y is a group representedby Formula (C)

(wherein R₂ is hydrogen; and l is 0 or 1), Z is —C(R₄)═C(R₅)(R₆); R₄ ishydrogen or fluorine; R₅ and R₆ are the same or different, and eachrepresents hydrogen, C₁-C₄ alkyl optionally having R₈, or a grouprepresented by Formula (D)

R₇ is hydrogen, hydroxymethyl, methyl, or 2-hydroxy-2-methyl-ethyl; R₈is —N(R_(x))(R_(y)); R₉ is C₁-C₄ alkyl, fluorine, or hydroxy; R_(x) andR_(y) are the same or different, and each represents hydrogen, C₁-C₄alkyl, C₃-C₁₀ cycloalkyl, or C₁-C₄ alkoxy-C₁-C₄ alkyl, m is 1, and n is0 or 1.

Even more preferable Compounds (I) are compounds represented by Formula(I) wherein R₁ is methyl or ethyl; X₁ and X₂ are such that (1) when X₂is N, X₁ is N or CH, and (2) when X₂ is CH, X₁ is CH; in Y, the divalentmoiety represented by

is pyrrolidinylene, azetidinylene, piperidinylene, or the divalentmoiety represented by

is phenylene;(a) when Y is a group represented by Formula (A)

(wherein R₂ is hydrogen, ethynyl, methoxycarbonyl, hydroxymethyl, or1,3,4-oxadiazolyl optionally having methyl; and l is 0 or 1), Z is—C(R₄)═C(R₅) (R₆) or —C≡C—R₇, (b) when Y is a group represented byFormula (C)

(wherein R₂ is hydrogen; and l is 1),Z is —C(R₄)═C(R₅) (R₆); R₄ is hydrogen; R₅ and R₆ are the same ordifferent, and each represents hydrogen, methyl having R₈, or themonovalent moiety represented by

of the group represented by Formula (D)

is pyrrolidinyl, piperidinyl, azetidinyl, piperazinyl, or morpholinyl;R₇ is hydrogen, hydroxymethyl, methyl, or 2-hydroxy-2-methyl-ethyl; R₈is —N(R_(x))(R_(y)); R₉ is methyl, fluorine, or hydroxy and; R_(x) andR_(y) are the same or different, and each represents hydrogen, methyl,ethyl, tert-butyl, isopropyl, cyclopropyl, cyclobutyl, or2-methoxyethyl; and m is 1, and n is 0 or 1.

Specific examples of preferable Compounds (I) include the following:

-   (1)    (S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one    (Compound of Example 2),-   (2)    (S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-yn-1-one    (Compound of Example 5),-   (3)    (S)-1-(3-(4-amino-3-((3,5-diethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one    (Compound of Example 8),-   (4)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)prop-2-en-1-one    (Compound of Example 9),-   (5)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-hydroxybut-2-yn-1-one    (Compound of Example 10),-   (6)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(dimethylamino)but-2-en-1-one    (Compound of Example 12),-   (7)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(cyclopropylamino)but-2-en-1-one    (Compound of Example 13),-   (8)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(isopropylamino)but-2-en-1-one    (Compound of Example 14),-   (9)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(ethyl(methyl)amino)but-2-en-1-one    (Compound of Example 15),-   (10)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(cyclobutylamino)but-2-en-1-one    (Compound of Example 16),-   (11)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(diethylamino)but-2-en-1-one    (Compound of Example 17),-   (12)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(tert-butylamino)but-2-en-1-one    (Compound of Example 18),-   (13)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(isopropyl(methyl)amino)but-2-en-1-one    (Compound of Example 19),-   (14)    1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(piperidin-1-yl)but-2-en-1-one    (Compound of Example 20),-   (15)    (S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(3-fluoropyrrolidin-1-yl)but-2-en-1-one    (Compound of Example 22), (16)    (R)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(3-fluoropyrrolidin-1-yl)but-2-en-1-one    (Compound of Example 23),-   (17)    1-((2S,4S)-4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-(hydroxymethyl)pyrrolidin-1-yl)prop-2-en-1-one    (Compound of Example 28),-   (18)    1-(2S,4S)-4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynylpyrrolidin-1-yl)prop-2-en-1-one    (Compound of Example 32),-   (19)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one    (Compound of Example 38),-   (20)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)prop-2-en-1-one    (Compound of Example 39),-   (21)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(pyrrolidin-1-yl)but-2-en-1-one    (Compound of Example 40),-   (22)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(4-hydroxypiperidin-1-yl)but-2-en-1-one    (Compound of Example 42),-   (23)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)but-2-yn-1-one    (Compound of Example 46),-   (24)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-hydroxy-4-methylpent-2-yn-1-one    (Compound of Example 47),-   (25)    1-((S)-3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-((S)-3-fluoropyrrolidin-1-yl)but-2-en-1-one    (Compound of Example 49),-   (26)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(piperidin-1-yl)but-2-en-1-one    (Compound of Example 50),-   (27)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)prop-2-en-1-one    (Compound of Example 51),-   (28)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(dimethylamino)but-2-en-1-one    (Compound of Example 52),-   (29)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(pyrrolidin-1-yl)but-2-en-1-one    (Compound of Example 53),-   (30)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(azetidin-1-yl)but-2-en-1-one    (Compound of Example 55),-   (31)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(ethyl(methyl)amino)but-2-en-1-one    (Compound of Example 56),-   (32)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(isopropylamino)but-2-en-1-one    (Compound of Example 57),-   (33)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(diethylamino)but-2-en-1-one    (Compound of Example 59),-   (34)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-((2-methoxyethyl)    (methyl)amino)but-2-en-1-one (Compound of Example 60),-   (35)    1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(4-hydroxypiperidin-1-yl)but-2-en-1-one    (Compound of Example 61),-   (36)    (S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(3-hydroxypyrrolidin-1-yl)but-2-en-1-one    (Compound of Example 62),-   (37)    (R)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(3-hydroxypyrrolidin-1-yl)but-2-en-1-one    (Compound of Example 63),-   (38)    (2S,4S)-methyl-1-acryloyl-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate    (Compound of Example 66),-   (39)    1-((2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(1,3,4-oxadiazol-2-yl)pyrrolidin-1-yl)prop-2-en-1-one    (Compound of Example 68), and-   (40)    (S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one    (Compound of Example 73).

Next, the method for producing Compound (I), which is the activeingredient of the antitumor agent according to the present invention,will be explained. Compound (I) can be produced, for example, by thefollowing production methods or by the methods described in Examples.However, the method for producing Compound (I) is not limited to thesereaction examples.

Production Method 1

(wherein P₁ is a protecting group of the amino group contained in Y; andR₁, X₁, X₂, Y, and Z are as defined above.)

(Step 1) In this step, the protected amino group of the compound ofFormula (II) is deprotected to produce the compound of Formula (III).The method for the deprotection can be performed according to a knownmethod, such as the method described in Protective Groups in OrganicSynthesis, T.W. Greene, John Wiley & Sons (1981); or methods similarthereto. An example of the protecting group is tert-butyloxycarbonyl. Ifa tert-butyloxycarbonyl group is used as a protecting group, thedeprotection is preferably performed under acidic conditions. Examplesof acids that can be used include hydrochloric acid, acetic acid,trifluoroacetic acid, sulfuric acid, methanesulfonic acid, tosic acid,and the like. Such an acid is preferably used in an amount of 1 to 100moles per mole of Compound (II).

Any solvent that does not adversely affect the reaction can be used.Examples thereof include alcohols (e.g., methanol), hydrocarbons (e.g.,benzene, toluene, and xylene), halogenated hydrocarbons (e.g., methylenechloride, chloroform, and 1,2-dichloroethane), nitriles (e.g.,acetonitrile), ethers (e.g., dimethoxyethane and tetrahydrofuran),aprotic polar solvents (e.g., N,N-dimethylformamide, dimethyl sulfoxide,and hexamethylphosphoramide), or a mixture thereof. The reaction time is0.1 to 100 hours, and preferably 0.5 to 24 hours. The reactiontemperature is 0 to 120° C., and preferably 0 to 90° C.

The thus-obtained compound of Formula (III) can be subjected to thesubsequent step after or without isolation and purification by knownseparation and purification means, such as concentration, vacuumconcentration, crystallization, solvent extraction, reprecipitation, andchromatography.

(Step 2) In this step, the compound of Formula (III) is amidated with acarboxylic acid represented by Z—COOH or with an acid halide representedby Z—C(═O)-L (wherein L is chlorine or bromine) to produce the compoundof Formula (I).

When a carboxylic acid represented by Z—COOH is used as an amidationreagent, the reaction is performed by using the carboxylic acid in anamount of 0.5 to 10 moles, and preferably 1 to 3 moles, per mole of thecompound of Formula (III) in the presence of a suitable condensingagent. The carboxylic acid may be a commercially available product, orcan be produced according to a known method.

Any reaction solvent that does not adversely affect the reaction can beused. Examples of preferable solvents include isopropanol, tert-butylalcohol, toluene, benzene, methylene chloride, chloroform,tetrahydrofuran, 1,4-dioxane, dimethylformamide, dimethylacetamide,N-methylpyrrolidinone, dimethyl sulfoxide, and mixed solvents thereof.The reaction temperature is usually −78 to 200° C., and preferably 0 to50° C. The reaction time is typically 5 minutes to 3 days, andpreferably 5 minutes to 10 hours.

Examples of the condensing agent include diphenylphosphoryl azide,N,N′-dicyclohexylcarbodiimide,benzotriazol-1-yloxy-trisdimethylaminophosphonium salts,4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmopholinium chloride,1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, a combination of1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and1-hydroxybenzotriazole, 2-chloro-1,3-dimethylimidazolinium chloride,O-(7-azabenzotriazo-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate, and the like.

A base can be optionally added for the reaction. Examples of usablebases include organic bases such as triethylamine,diisopropylethylamine, pyridine, lutidine, collidine,4-dimethylaminopyridine, potassium tert-butyrate, sodium tert-butyrate,sodium methoxide, sodium ethoxide, lithium hexamethyldisilazide, sodiumhexamethyldisilazide, potassium hexamethyldisilazide, and butyl lithium;and inorganic bases such as sodium hydrogen carbonate, sodium carbonate,potassium carbonate, cesium carbonate, sodium hydroxide, and sodiumhydride. Such a base is added in an amount of 1 to 100 moles, andpreferably 1 to 10 moles, per mole of the compound of Formula (III).

When an acid halide represented by Z—C(═O)-L (wherein L is chlorine orbromine) is used as an amidation reagent, the acid halide is used in anamount of 0.5 to 5 moles, and preferably 0.9 to 1.1 moles, per mole ofthe compound of Formula (III). The acid halide may be a commerciallyavailable product, or can be produced according to a known method.

Any reaction solvent that does not adversely affect the reaction can beused. Examples of preferable solvents include toluene, benzene,methylene chloride, chloroform, tetrahydrofuran, 1,4-dioxane,dimethylformamide, dimethylacetamide, N-methylpyrrolidinone, and mixedsolvents thereof. The reaction temperature is typically −78 to 200° C.,and preferably −20 to 50° C. The reaction time is typically 5 minutes to3 days, and preferably 5 minutes to 10 hours.

A base can be optionally added for the reaction. Examples of usablebases include organic bases such as triethylamine,diisopropylethylamine, pyridine, lutidine, collidine,4-dimethylaminopyridine, potassium tert-butyrate, sodium tert-butyrate,sodium methoxide, sodium ethoxide, lithium hexamethyldisilazide, sodiumhexamethyldisilazide, potassium hexamethyldisilazide, and butyl lithium;and inorganic bases such as sodium hydrogen carbonate, sodium carbonate,potassium carbonate, cesium carbonate, sodium hydroxide, and sodiumhydride. Such a base is added in an amount of 1 to 100 moles, preferably1 to 10 moles, per mole of the compound of Formula (III).

The thus-obtained compound of Formula (I) can be isolated and purifiedby known separation and purification means, such as concentration,vacuum concentration, crystallization, solvent extraction,reprecipitation, and chromatography.

Among Compounds (I), compound of Formula (I′) or (I″) can also beproduced by production method 2 using, for example, Compound (III)obtained in step 1 of production method 1 as a starting compound, andusing a specific amine.

Production Method 2

(wherein L₁ and L₂ are halogen; H of Y—H is hydrogen directly bound to anitrogen atom; and X₁, X₂, Y, R_(x), R_(y), R₁,

m, and n are as defined above.)

(Step 3) In this step, the compound of Formula (III) is amidated with anacid halide represented by Formula (IV) to produce the compound ofFormula (V).

Examples of halogen atoms represented by L₁ or L₂ in Formula (IV)include bromine and chlorine. The compound represented by Formula (IV)may be a commercially available product, or can be produced according toa known method.

The compound of Formula (IV) is used in an amount of 0.5 to 5 moles, andpreferably 0.9 to 1.1 moles, per mole of Compound (III).

A base can be optionally added for the reaction. Examples of usablebases include organic bases such as triethylamine,diisopropylethylamine, pyridine, lutidine, collidine,4-dimethylaminopyridine, potassium tert-butyrate, sodium tert-butyrate,sodium methoxide, sodium ethoxide, lithium hexamethyldisilazide, sodiumhexamethyldisilazide, potassium hexamethyldisilazide, and butyl lithium;and inorganic bases such as sodium hydrogen carbonate, sodium carbonate,potassium carbonate, cesium carbonate, sodium hydroxide, and sodiumhydride. Such a base can be added in an amount of 1 to 100 moles, andpreferably 1 to 10 moles, per mole of the compound of Formula (III).

Any reaction solvent that does not adversely affect the reaction can beused. Examples of preferable reaction solvents include toluene, benzene,methylene chloride, chloroform, tetrahydrofuran, 1,4-dioxane,dimethylformamide, dimethylacetamide, N-methylpyrrolidinone, and mixedsolvents thereof. The reaction temperature is typically −78 to 200° C.,and preferably 0 to 50° C. The reaction time is typically 5 minutes to 3days, and preferably 5 minutes to 10 hours.

The thus-obtained compound of Formula (V) can be subjected to thesubsequent step after or without isolation and purification by knownseparation and purification means, such as concentration, vacuumconcentration, crystallization, solvent extraction, reprecipitation, andchromatography.

(Step 4) In this step, the compound of Formula (V) is alkylated with anamine represented by Formula (VI) or (VI′) to produce the compound ofFormula (I′) or (I″).

The compound of Formula (VI) or (VI′) can be used in an amount of 1 to20 moles, and preferably 1 to 10 moles, per mole of the compound ofFormula (V).

Further, a base can optionally be added for the reaction. Examples ofsuch bases include organic bases such as triethylamine,diisopropylethylamine, pyridine, lutidine, collidine,4-dimethylaminopyridine, potassium tert-butyrate, sodium tert-butyrate,sodium methoxide, sodium ethoxide, lithium hexamethyldisilazide, sodiumhexamethyldisilazide, potassium hexamethyldisilazide, and butyl lithium;and inorganic bases such as sodium hydrogen carbonate, sodium carbonate,potassium carbonate, cesium carbonate, sodium hydroxide, and sodiumhydride. Such a base can be added in an amount of 1 to 100 moles, andpreferably 1 to 20 moles, per mole of the compound of Formula (V).

Any reaction solvent that does not adversely affect the reaction can beused. For example, N,N-dimethylformamide, dimethylacetamide, dimethylsulfoxide, tetrahydrofuran, 1,4-dioxane, N-methylpyrrolidin-2-one,acetonitrile, and the like can be used singly, or as a mixture. Thereaction time is 0.1 to 100 hours, and preferably 0.5 to 24 hours. Thereaction temperature is 0° C. to the boiling temperature of the solvent,and preferably 0 to 100° C.

The thus-obtained compound of Formula (I′) or (I″) can be isolated andpurified by known separation and purification means, such asconcentration, vacuum concentration, crystallization, solventextraction, reprecipitation, and chromatography.

The compound of Formula (II) used for producing Compound (I) can beproduced, for example, by production method 3 or 4.

Production Method 3

(wherein L₃ and L₄ are leaving groups; and R₁, X₁, X₂, Y, and P₁ are asdefined above.)

(Step 5) In this step, the compound of Formula (VII) is subjected to acoupling (Sonogashira) reaction with the compound of Formula (VIII) toproduce the compound of Formula (IX). This step can be performedaccording to a generally known method (see, for example, ChemicalReviews, vol. 107, p. 874, 2007), for example, in the presence of atransition metal catalyst and a base in a solvent that does notadversely affect the reaction.

In Formula (VII), the leaving group represented by L₃ is bromine oriodine. The compound of Formula (VII) may be a commercially availableproduct, or can be produced by a known method.

In this step, the compound of Formula (VIII) can be used in an amount of1 to 10 moles, and preferably 1 to 3 moles, per mole of the compound ofFormula (VII).

Examples of transition metal catalysts that can be used in this stepinclude palladium catalysts (e.g., palladium acetate,tris(dibenzylideneacetone)dipalladium, and1,1′-bis(diphenylphosphino)ferrocene-palladium(II) dichloridedichloromethane complex). If necessary, a ligand (e.g.,triphenylphosphine and tri-tert-butylphosphine) can be added and acopper reagent (e.g., copper iodide and copper acetate) can be used as acocatalyst. The amount of the transition metal catalyst used may varydepending on the type of catalyst. The transition metal catalyst istypically used in an amount of 0.0001 to 1 mole, and preferably 0.01 to0.5 moles, per mole of the compound of Formula (VII). The amount of theligand used is typically 0.0001 to 4 moles, and preferably 0.01 to 2moles, per mole of the compound of Formula (VII). The amount of thecocatalyst used is typically 0.0001 to 4 moles, and preferably 0.01 to 2moles, per mole of the compound of Formula (VII).

A base may optionally be added for the reaction. Examples of usablebases include organic bases such as triethylamine,diisopropylethylamine, pyridine, lutidine, collidine,4-dimethylaminopyridine, potassium tert-butyrate, sodium tert-butyrate,sodium methoxide, sodium ethoxide, lithium hexamethyldisilazide, sodiumhexamethyldisilazide, potassium hexamethyldisilazide, and butyl lithium;and inorganic bases such as sodium hydrogen carbonate, sodium carbonate,potassium carbonate, cesium carbonate, sodium hydroxide, and sodiumhydride. Among them, organic bases such as triethylamine anddiisopropylethylamine are preferable. The amount of the base used istypically 0.1 to 50 moles, preferably 1 to 20 moles, per mole of thecompound of Formula (VII).

Any reaction solvent that does not adversely affect the reaction can beused. Examples of usable solvents include hydrocarbons (e.g., benzene,toluene, and xylene), nitriles (e.g., acetonitrile), ethers (e.g.,dimethoxyethane, tetrahydrofuran, and 1,4-dioxane), alcohols (e.g.,methanol and ethanol), aprotic polar solvents (e.g., dimethylformamide,dimethyl sulfoxide, and hexamethylphosphoramide), water, and mixturesthereof. The reaction time is 0.1 to 100 hours, and preferably 0.5 to 24hours. The reaction temperature is 0° C. to the boiling temperature ofthe solvent, and preferably 0 to 150° C.

The thus-obtained compound of Formula (IX) can be subjected to thesubsequent step after or without isolation and purification by knownseparation and purification means, such as concentration, vacuumconcentration, crystallization, solvent extraction, reprecipitation, andchromatography.

(Step 6) In this step, the compound of Formula (IX) is used with thecompound of Formula (X) or (XI) to produce the compound of Formula (II).

When the compound of Formula (X) is used as an alkylating reagent, thecompound of Formula (II) can be produced in the presence of a base. InFormula (X), L₄ may be a leaving group, such as chlorine, bromine,iodine, a methanesulfonic acid ester, or a p-toluenesulfonic acid ester.The alkylating reagent may be a commercially available product, or canbe produced according to a known method. The compound of Formula (X) canbe used in an amount of 1 to 10 moles, preferably 1 to 5 moles, per moleof the compound of Formula (IX).

Examples of usable bases include inorganic bases such as sodium hydrogencarbonate, sodium carbonate, potassium carbonate, cesium carbonate,cesium hydroxide, sodium hydride, and potassium hydride; and organicamines such as trimethylamine, triethylamine, tripropylamine,diisopropylethylamine, N-methylmorpholine, pyridine,4-(N,N-dimethylamino)pyridine, lutidine, and collidine. Such a base canbe used in an amount of 1 to 100 moles, preferably 2 to 10 moles, permole of the compound of Formula (IX).

As the solvent, N,N-dimethylformamide, N,N-dimethylacetamide, dimethylsulfoxide, tetrahydrofuran, 1,4-dioxane, N-methylpyrrolidin-2-one,acetonitrile, and the like can be used singly, or as a mixture. Thereaction time is 0.1 to 100 hours, and preferably 0.5 to 24 hours. Thereaction temperature is 0° C. to the boiling temperature of the solvent,and preferably 0 to 100° C.

When the compound of Formula (XI) is used as an alkylating reagent, thecompound of Formula (II) can be produced by using a Mitsunobu reaction.This step can be performed according to a generally known method (see,for example, Chemical Reviews, Vol. 109, p. 2551, 2009), for example, inthe presence of Mitsunobu reagents and a phosphine reagent in a solventthat does not adversely affect the reaction. This step is performedusing the compound of Formula (XI) in an amount of 1 to 10 moles permole of the compound of Formula (IX).

Examples of Mitsunobu reagents include diethyl azodicarboxylate,diisopropyl azodicarboxylate, and the like. Such Mitsunobu reagents areused in an amount of 1 to 10 moles, and preferably 1 to 5 moles, permole of the compound of Formula (IX).

Examples of phosphine reagents include triphenylphosphine,tributylphosphine, and the like. Such a phosphine reagent is used in anamount of 1 to 10 moles, and preferably 1 to 5 moles, per mole of thecompound of Formula (IX).

Any reaction solvent that does not adversely affect the reaction can beused. Examples of preferable reaction solvents include toluene,benzenetetrahydrofuran, 1,4-dioxane, dimethylformamide,dimethylacetamide, N-methylpyrrolidinone, dimethyl sulfoxide, and mixedsolvents thereof.

The reaction temperature is typically −78 to 200° C., and preferably 0to 50° C. The reaction time is typically 5 minutes to 3 days, andpreferably 10 minutes to 10 hours.

The thus-obtained compound of Formula (II) can be utilized after orwithout isolation and purification by known separation and purificationmeans, such as concentration, vacuum concentration, crystallization,solvent extraction, reprecipitation, and chromatography, to produceCompound (I).

Production Method 4

(wherein L₃, L₄, R₁, X₁, X₂, Y, and P₁ are as defined above.)

(Step 7) This step can be performed in a manner similar to step 6.

(Step 8) This step can be performed in a manner similar to step 5.

The compound of Formula (XII) used in the production of Compound (I) canalso be produced, for example, by production method 5.

Production Method 5

(wherein L₃, L₄, X₁, X₂, Y, and P₁ are as defined above.)

(Step 9) This step can be performed in a manner similar to step 6.

(Step 10) In this step, the compound of Formula (XIV) is reacted withammonia or a salt thereof to produce the compound of Formula (XII).

The ammonia or a salt thereof is typically used in an equimolar toexcessive molar amount per mole of the compound of Formula (XIII) inthis step. Any reaction solvent that does not adversely affect thereaction can be used. Examples of preferable reaction solvents includewater, methanol, ethanol, isopropanol, tert-butyl alcohol,tetrahydrofuran, 1,4-dioxane, dimethylformamide, N-methylpyrrolidone,dimethyl sulfoxide, and mixed solvents thereof.

The reaction temperature is typically 0 to 200° C., and preferably roomtemperature to 150° C. The reaction time is typically 5 minutes to 7days, and preferably 30 minutes to 24 hours.

The thus-obtained compound of Formula (XIV) can be subjected to thesubsequent step 8 after or without isolation and purification by knownseparation and purification means, such as concentration, vacuumconcentration, crystallization, solvent extraction, reprecipitation, andchromatography.

The compound of Formula (IX) used in the production of Compound (I) canalso be produced, for example, by production method 6.

Production Method 6

(wherein L₃, X₁, and X₂ are as defined above, and SEM istrimethylsilylethoxymethyl.)

(Step 11) In this step, the compound of Formula (XIII) is reacted withSEMCl (trimethylsilylethoxymethylchloride) in the presence of a base toproduce the compound of Formula (XV). The compound of Formula (XIII) maybe a commercially available product, or can be produced according to aknown method.

SEMCl is typically used in an equimolar to excessive molar amount permole of the compound of Formula (XIII) in this step. Any reactionsolvent that does not adversely affect the reaction can be used.Examples of preferable reaction solvents include tetrahydrofuran,1,4-dioxane, chloroform, methylene chloride, dimethylformamide,N-methylpyrrolidone, and mixed solvents thereof.

Examples of usable bases include organic bases such as triethylamine,diisopropylethylamine, pyridine, and 4-dimethylaminopyridine; andinorganic bases such as sodium hydrogen carbonate, sodium carbonate,potassium carbonate, cesium carbonate, sodium hydroxide, sodium hydride,and potassium tert-butyrate.

Such a base is typically used in an equimolar to excessive molar amount,and preferably 1 to 3 moles, per mole of the compound of Formula (XIII).

The reaction temperature is typically −78 to 50° C., and preferably 0°C. to room temperature. The reaction time is typically 5 minutes to 7days, and preferably 10 minutes to 24 hours.

The thus-obtained compound of Formula (XV) can be subjected to thesubsequent step after or without isolation and purification by knownseparation and purification means, such as concentration, vacuumconcentration, crystallization, solvent extraction, reprecipitation, andchromatography.

(Step 12) This step can be performed in a manner similar to step 10.

The thus-obtained compound of Formula (XVI) can be subjected to thesubsequent step after or without isolation and purification by knownseparation and purification means, such as concentration, vacuumconcentration, crystallization, solvent extraction, reprecipitation, andchromatography.

(Step 13) This step can be performed in a manner similar to step 5.

(Step 14) In this step, the compound of Formula (XVII) is deprotectedunder acidic conditions to produce the compound of Formula (IX). Thedeprotection can be performed by a known method, such as the methoddescribed in Protective Groups in Organic Synthesis, T.W. Greene, JohnWiley & Sons (1981); or a method similar thereto.

Examples of usable acids include hydrochloric acid, acetic acid,trifluoroacetic acid, sulfuric acid, methanesulfonic acid, tosic acid,and the like. Such an acid is used in an amount of 1 to 100 moles permole of the compound of Formula (XVII).

Any solvent that does not adversely affect the reaction can be used.Examples of usable solvents include alcohols (e.g., methanol),hydrocarbons (e.g., benzene, toluene, and xylene), halogenatedhydrocarbons (e.g., methylene chloride, chloroform, and1,2-dichloroethane), nitriles (e.g., acetonitrile), ethers (e.g.,dimethoxyethane, tetrahydrofuran, and 1,4-dioxane), aprotic polarsolvents (e.g., N,N-dimethylformamide, dimethyl sulfoxide, andhexamethylphosphoramide), and mixtures thereof. The reaction time is 0.1to 100 hours, and preferably 0.5 to 24 hours. The reaction temperatureis 0° C. to the boiling temperature of the solvent, and preferably 0 to100° C.

The thus-obtained compound of Formula (IX) can be used in step 6 afteror without isolation and purification by known separation andpurification means, such as concentration, vacuum concentration,crystallization, solvent extraction, reprecipitation, andchromatography.

In the above production methods 1 to 6, for functional groups having anactive proton, such as amino, imino, hydroxy, carboxy, and amide groups,and indole, protected reagents can be used or a protecting group isintroduced into such a functional group according to a usual method, andthen the protecting group can be removed in an appropriate step in eachproduction method.

The “protecting group of an amino group or protecting group of an iminogroup” is not particularly limited insofar as it has a protectingfunction. Examples of such protecting groups include aralkyl groups suchas benzyl, p-methoxybenzyl, 3,4-dimethoxybenzyl, o-nitrobenzyl,p-nitrobenzyl, benzhydryl, trityl, and cumyl; lower alkanoyl groups suchas formyl, acetyl, propionyl, butyryl, pivaloyl, trifluoroacetyl, andtrichloroacetyl; benzoyl; arylalkanoyl groups such as phenylacetyl andphenoxyacetyl; lower alkoxycarbonyl groups such as methoxycarbonyl,ethoxycarbonyl, propyloxycarbonyl, and tert-butoxycarbonyl;aralkyloxycarbonyl groups such as p-nitrobenzyloxycarbonyl andphenethyloxycarbonyl; lower alkylsilyl groups such as trimethylsilyl andtert-butyldimethylsilyl; tetrahydropyranyl; trimethylsilylethoxymethyl;lower alkylsulfonyl groups such as methylsulfonyl, ethylsulfonyl, andtert-butylsulfonyl; lower alkylsulfinyl groups such astert-butylsulfinyl; arylsulfonyl groups such as benzenesulfonyl andtoluenesulfonyl; and imido groups such as phthalimido. In particular,trifluoroacetyl, acetyl, tert-butoxycarbonyl, benzyloxycarbonyl,trimethylsilylethoxymethyl, cumyl, and the like are preferable.

The “protecting group of a hydroxy group” is not particularly limitedinsofar as it has a protecting function. Examples of such protectinggroups include lower alkyl groups such as methyl, ethyl, propyl,isopropyl, and tert-butyl; lower alkylsilyl groups such astrimethylsilyl and tert-butyldimethylsilyl; lower alkoxymethyl groupssuch as methoxymethyl and 2-methoxyethoxymethyl; tetrahydropyranyl;trimethylsilylethoxymethyl; aralkyl groups such as benzyl,p-methoxybenzyl, 2,3-dimethoxybenzyl, o-nitrobenzyl, p-nitrobenzyl, andtrityl; and acyl groups such as formyl, acetyl, and trifluoroacetyl. Inparticular, methyl, methoxymethyl, tetrahydropyranyl,trimethylsilylethoxymethyl, tert-butyldimethylsilyl, acetyl, and thelike are preferable.

The “protecting group of a carboxy group” is not particularly limitedinsofar as it has a protecting function. Examples of such protectinggroups include lower alkyl groups such as methyl, ethyl, propyl,isopropyl, and tert-butyl; halo-lower-alkyl groups such as2,2,2-trichloroethyl; lower alkenyl groups such as allyl;trimethylsilylethoxymethyl; and aralkyl groups such as benzyl,p-methoxybenzyl, p-nitrobenzyl, benzhydryl, and trityl. In particular,methyl, ethyl, tert-butyl, allyl, benzyl, p-methoxybenzyl,trimethylsilylethoxymethyl, and the like are preferable.

The “protecting group of a carbonyl group” is not particularly limitedinsofar as it has a protecting function. Examples of such protectinggroups include ketals and acetals, such as ethylene ketal, trimethyleneketal, dimethyl ketal, ethylene acetal, trimethylene acetal, anddimethyl acetal.

The method for removing such a protecting group may vary depending onthe type of protecting group, stability of the desired compound (I),etc. For example, the following methods can be used: solvolysis using anacid or a base according to the method disclosed in a publication(Protective Groups in Organic Synthesis, third edition, T.W. Green, JohnWiley & Sons (1999)) or a method similar thereto, i.e., a reactionmethod using, for example, 0.01 moles or a large excess of an acid,preferably trifluoroacetic acid, formic acid, or hydrochloric acid, oran equimolar amount to a large excess of a base, preferably potassiumhydroxide or calcium hydroxide; chemical reduction using a metal hydridecomplex or the like; or catalytic reduction using a palladium-carboncatalyst, Raney nickel catalyst, or the like.

Compound (I) can easily be isolated and purified by usual isolation andpurification means. Examples of such means include solvent extraction,recrystallization, preparative reversed-phase high-performance liquidchromatography, column chromatography, preparative thin-layerchromatography, and the like.

When Compound (I) has isomers such as optical isomers, stereoisomers,positional isomers, and rotational isomers, any of the isomers andmixtures thereof are included within the scope of Compound (I). Forexample, when Compound (I) has optical isomers, optical isomersseparated from a racemic mixture are also included within the scope ofCompound (I). Each of these isomers can be obtained as a single compoundby known synthesis and separation means (e.g., concentration, solventextraction, column chromatography, and recrystallization).

Compound (I) or a salt thereof may be crystalline. A single crystal formthereof and a polymorphic mixture thereof are both included within thescope of Compound (I) or a salt thereof. These crystals can be producedby crystallization according to a crystallization method known per se inthe art. Compound (I) or a salt thereof may be a solvate (e.g., ahydrate) or a non-solvate. Any of such forms are included within thescope of Compound (I) or a salt thereof. Compounds labeled with anisotope (such as ³H, ¹⁴C, ³⁵S, or ¹²⁵I) are also included within thescope of Compound (I) or a salt thereof.

A prodrug of Compound (I) or a salt thereof refers to a compound thatcan be converted to Compound (I) or a salt thereof through a reactionwith an enzyme, gastric acid, or the like under physiological conditionsin vivo, i.e., a compound that can be converted to Compound (I) or asalt thereof by enzymatic oxidation, reduction, hydrolysis, or the like;or a compound that can be converted to Compound (I) or a salt thereof byhydrolysis with gastric acid or the like. Further, the prodrug ofCompound (I) or a salt thereof may be compounds that can be converted toCompound (I) or a salt thereof under physiological conditions, such asthose described in “Iyakuhin no Kaihatsu [Development ofPharmaceuticals],” Vol. 7, Molecular Design, published in 1990 byHirokawa Shoten Co., pp. 163-198.

The salt of Compound (I) refers to a common salt used in the field oforganic chemistry. Examples of such salts include base addition salts tocarboxyl when the compound has carboxyl, and acid addition salts to anamino or basic heterocyclic group when the compound has an amino orbasic heterocyclic group.

Examples of base addition salts include alkali metal salts such assodium salts and potassium salts; alkaline earth metal salts such ascalcium salts and magnesium salts; ammonium salts; and organic aminesalts such as trimethylamine salts, triethylamine salts,dicyclohexylamine salts, ethanolamine salts, diethanolamine salts,triethanolamine salts, procaine salts, and N,N′-dibenzylethylenediaminesalts.

Examples of acid addition salts include inorganic acid salts such ashydrochlorides, sulfates, nitrates, phosphates, and perchlorates;organic acid salts such as acetates, formates, maleates, fumarates,tartrates, citrates, ascorbates, and trifluoroacetates; and sulfonatessuch as methanesulfonates, isethionates, benzenesulfonates, andp-toluenesulfonates.

Compound (I) or a salt thereof has excellent FGFR inhibitory activity,and is useful as an antitumor agent. Further, Compound (I) or a saltthereof has excellent selectivity toward FGFR, and has advantageouslyfewer side effects caused by other kinases. Although the target canceris not particularly limited, examples thereof include head and neckcancer, esophagus cancer, gastric cancer, colon cancer, rectum cancer,liver cancer, gallbladder cancer, cholangiocarcinoma, biliary tractcancer, pancreatic cancer, lung cancer, breast cancer, ovarian cancer,cervical cancer, endometrial cancer, renal cancer, bladder cancer,prostate cancer, testicular tumor, osteosarcoma, soft-tissue sarcoma,blood cancer, multiple myeloma, skin cancer, brain tumor, andmesothelioma. Preferably, the target cancer is blood cancers such asB-cell lymphoma, chronic lymphocytic leukemia, peripheral T-celllymphoma, myelodysplastic syndrome, acute myeloid leukemia, and acutelymphocytic leukemia.

When Compound (I) or a salt thereof is used as a pharmaceuticalpreparation, a pharmaceutical carrier can be added, if required, therebyforming a suitable dosage form according to prevention and treatmentpurposes. Examples of the dosage form include oral preparations,injections, suppositories, ointments, patches, and the like. Of these,oral preparations are preferable. Such dosage forms can be formed bymethods conventionally known to persons skilled in the art.

As the pharmaceutical carrier, various conventional organic or inorganiccarrier materials used as preparation materials may be blended as anexcipient, binder, disintegrant, lubricant, or colorant in solidpreparations; or as a solvent, solubilizing agent, suspending agent,isotonizing agent, buffer, or soothing agent in liquid preparations.Moreover, pharmaceutical preparation additives, such as antiseptics,antioxidants, colorants, sweeteners, and stabilizers, may also be used,if required.

Oral solid preparations can be prepared as follows. An excipient,optionally together with a binder, disintegrant, lubricant, colorant,taste-masking or flavoring agent, etc., is added to Compound (I) or asalt thereof to produce tablets, coated tablets, granules, powders,capsules, or the like, using an ordinary method.

When an injection agent is prepared, a pH adjuster, buffer, stabilizer,isotonizing agent, local anesthetic, etc., may be added to Compound (I)or a salt thereof; and the mixture may be processed into a subcutaneous,intramuscular, or intravenous injection according to an ordinary method.

The amount of Compound (I) or a salt thereof to be contained in such adosage unit form varies depending on the condition of the patient, thedosage form, etc. The desirable amount in dosage unit form is generally0.05 to 1,000 mg in the case of an oral preparation, 0.01 to 500 mg inthe case of an injection, and 1 to 1,000 mg in the case of asuppository.

Moreover, the daily dose of the medicine having the above-describeddosage form may vary depending on the condition, body weight, age, andsex of a patient, etc., and cannot be generalized. Usually, the dailydose is preferably 0.05 to 5000 mg per adult (body weight: 50 kg) perday, and more preferably 0.1 to 1000 mg per adult (body weight: 50 kg)per day. Such a dose of the medicine is preferably administered in onedose, or in two to three divided doses, per day.

EXAMPLES

The present invention is explained in more detail below with referenceto Examples and Test Examples; however, the scope of the presentinvention is not limited to these Examples.

In the Examples, commercially available reagents were used, unlessotherwise specified. Purif-Pack (registered trademark) SI, produced byMoritex Corp.; KP-Sil (registered trademark) Silica prepacked column,produced by Biotage; or HP-Sil (registered trademark) Silica prepackedcolumn, produced by Biotage was used as the silica gel columnchromatography. Purif-Pack (registered trademark) NH, produced byMoritex Corp; or KP-NH (registered trademark) prepacked column, producedby Biotage was used as the basic silica gel column chromatography.Kieselgel TM 60F 254, Art. 5744, produced by Merck, or NH₂ Silica Gel60F254 Plate, produced by Wako, was used as the preparative thin-layerchromatography. NMR spectrum was measured by using AL400 (400 MHz;produced by JEOL), Mercury 400 (400 MHz; produced by AgilentTechnologies, Inc.) spectrometer, or Inova 400 (400 MHz; produced byAgilent Technologies, Inc.) model spectrometer equipped with an OMNMRprobe (produced by Protasis). When its deuterated solvent containstetramethylsilane, the tetramethylsilane was used as the internalreference; and when tetramethylsilane is not contained, an NMR solventwas used as the reference. All the delta values are shown by ppm. Themicrowave reaction was performed using Discover S-class, produced by CEMCorporation.

The LCMS spectrum was measured using an Acquity SQD (quadrupole),produced by Waters Corporation, under the following conditions.

Column: YMC-Triart C18, 2.0×50 mm, 1.9 μm (produced by YMC)

MS detection: ESI positive

UV detection: 254 and 210 nm

Column flow rate: 0.5 mL/min

Mobile phase: Water/acetonitrile (0.1% formic acid)

Injection volume: 1 μL

TABLE 1 Gradient Time (min) Water Acetonitrile 0 95 5 0.1 95 5 2.1 5 953.0 STOP

Preparative reversed-phase HPLC purification was performed using apreparative separation system available from

Waters Corporation, under the following conditions.

Column: Connected YMC-Actus Triart C18, 20×50 mm, 5 μm

(produced by YMC) and YMC-Actus Triart C18, 20×10 mm, 5 μm

(produced by YMC).

UV detection: 254 nm

MS detection: ESI positive

Column flow rate: 25 mL/min

Mobile phase: Water/acetonitrile (0.1% formic acid)

Injection volume: 0.1 to 0.5 mL

Each symbol stands for the following.

s: Singlet

d: Doublet

t: Triplet

q: Quartet

dd: Double Doublet

dt: Double Triplet

td: Triple Doublet

tt: Triple Triplet

ddd: Double Double Doublet

ddt: Double Double Triplet

dtd: Double Triple Doublet

tdd: Triple Double Doublet

m: Multiplet

br: Broad

brs: Broad Singlet

DMSO-d₆: Deuterated dimethyl sulfoxide

CDCl₃: Deuterated chloroform

CD₃OD: Deuterated methanol

THF: Tetrahydrofuran

DMF: N,N-dimethylformamide

NMP: 1-Methyl-2-pyrrolidinone

DMSO: Dimethyl sulfoxide

TFA: Trifluoroacetic acid

SEMCl: 2-(Trimethylsilyl)ethoxymethyl chloride

PdCl₂ (dppf) CH₂Cl₂: 1,1′-bis(diphenylphosphino) ferrocene-palladium(II) dichloride-dichloromethane complex

WSC: 1-(3-Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride

HOBt: 1-Hydroxybenzotriazole monohydrate

HATU: O-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluroniumhexafluorophosphate

HBTU: O-benzotriazol-N,N,N′,N′-tetramethyluronium hexafluorophosphate

DIAD: Diisopropyl azodicarboxylate

TBAF: Tetrabutylammoniumfluolide

DIPEA: Diisopropylethylamine

BoC₂O: Di-tert-butyl dicarbonate

DMAP: Dimethylaminopyridine

Example 1 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one(Compound of Example 1) (Step 1) Synthesis of3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

PdCl₂ (dppf) CH₂Cl₂ (163 mg) was added to a mixture of3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (520 mg) synthesized by themethod disclosed in WO 2007/126841, 1-ethynyl-3,5-dimethoxybenzene (504mg), copper (I) iodide (57.3 mg), and triethylamine (0.56 ml) in DMF (10ml). After nitrogen purging, the resulting mixture was stirred at 90° C.for 6 hours. Chloroform and water were added to the reaction mixture toseparate the organic layer. After being washed with a saturated sodiumchloride solution, the organic layer was dried over anhydrous sodiumsulfate, and the solvent was then distilled off under reduced pressure.The resulting residue was purified by basic silica gel columnchromatography (developing solvent: chloroform/methanol) to obtain thetitle compound as a dark-brown solid (120 mg). Physical properties: m/z[M+H]⁺ 296.0

(Step 2) Synthesis of tert-butyl3-(methylsulfonyloxy)pyrrolidine-1-carboxylate

N-Boc-3-pyrrolidinol (1000 mg) was dissolved in chloroform (20 ml).Triethylamine (1.15 ml) and methanesulfonyl chloride (498 μl) were addedthereto at 0° C. After stirring at room temperature for 1.0 hour, ethylacetate and water were added thereto to separate the organic layer.After being washed with a saturated aqueous sodium bicarbonate solution,a saturated aqueous ammonium chloride solution and water, the organiclayer was dried over anhydrous sodium sulfate. The solvent was distilledoff under reduced pressure to obtain the title compound as a colorless,oily compound (1.2 g). Physical properties: m/z [M+H]⁺ 266.1

(Step 3) Synthesis of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate

A suspension of3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (62mg) obtained in Step 1, tert-butyl3-(methylsulfonyloxy)pyrrolidine-1-carboxylate (217 mg) obtained in Step2, and potassium carbonate (221 mg) in DMF (2.0 ml) was stirred at 70°C. for 1 hour. Ethyl acetate and water were added thereto to separatethe organic layer. The organic layer was dried over anhydrous sodiumsulfate, and the solvent was distilled off under reduced pressure. Theresulting residue was purified by basic silica gel column chromatography(developing solvent: hexane/ethyl acetate) to obtain the title compoundas a light-yellow, amorphous substance (36.2 mg). Physical properties:m/z [M+H]⁺ 465.1

(Step 4) Synthesis of Compound of Example 1

4N-Hydrochloric acid/1,4-dioxane (4 ml) was added to the tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate(32 mg) obtained in Step 3, and the mixture was stirred at roomtemperature for 1.5 hours. After distilling the solvent of the resultingreaction mixture off under reduced pressure, toluene azeotropicdistillation was subsequently performed to obtain a crude product of3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine(32 mg). Chloroform (2.0 ml) and triethylamine (20 μl) were added to aportion of the resulting crude product (12 mg). After cooling to 0° C.,acrylic chloride (2.3 μl) dissolved in chloroform (100 μl) was addedthereto, and the mixture was stirred at room temperature for 10 minutes.After halting the reaction using a saturated aqueous sodium bicarbonatesolution, the resulting product was extracted with ethyl acetate. Afterdrying the result over anhydrous sodium sulfate, the solvent wasdistilled off under reduced pressure. The resulting residue was purifiedby silica gel column chromatography (developing solvent: ethylacetate/methanol) to obtain the title compound as a white solid (6.5mg). Table 1 shows the physical properties thereof.

Example 2 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one(Compound of Example 2) (Step 1) Synthesis of (R)-tert-butyl3-(methylsulfonyloxy)pyrrolidine-1-carboxylate

(R)—N-Boc-3-pyrrolidinol (935 mg) was dissolved in chloroform (15 ml),and triethylamine (1.04 ml) and methanesulfonyl chloride (467 μl) wereadded thereto at 0° C. After stirring at room temperature for 1.5 hours,ethyl acetate and water were added thereto to separate the organiclayer. After being washed with a saturated aqueous sodium bicarbonatesolution, a saturated aqueous ammonium chloride solution and water, theorganic layer was dried over anhydrous sodium sulfate, and the solventwas distilled off under reduced pressure to obtain the title compound asa colorless, oily compound (1.1 g). Physical properties: m/z [M+H]⁺266.1

(Step 2) Synthesis of (S)-tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate

A suspension of3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine(334 mg) obtained in Example 1 (Step 1), (R)-tert-butyl3-(methylsulfonyloxy)pyrrolidine-1-carboxylate (379 mg) obtained in(Step 1) above, and potassium carbonate (391 mg) in DMF (4.0 ml) wasstirred at 70° C. for 3 hours. Ethyl acetate and water were addedthereto to separate the organic layer. The organic layer was dried overanhydrous sodium sulfate, and the solvent was then distilled off underreduced pressure. The resulting residue was purified by basic silica gelcolumn chromatography (developing solvent: hexane/ethyl acetate) toobtain the title compound as a light-yellow, amorphous substance (149mg). Physical properties: m/z [M+H]⁺ 465.1

(Step 3) Synthesis of Compound of Example 2

In accordance with Example 1 (Step 4), except that (S)-tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylateobtained in (Step 2) above was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,a crude product of(S)-3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-aminewas obtained by removing a Boc group under acidic conditions.Thereafter, amidation was conducted to obtain the title compound as awhite solid. Table 1 shows the physical properties thereof.

Example 3 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)but-2-en-1-one(Compound of Example 3)

In accordance with Example 1 (Step 4), except that 2-butenoyl chloridewas used in place of acrylic chloride, the title compound was obtainedas a white solid. Table 1 shows the physical properties thereof.

Example 4 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Compound of Example 4)

The crude product (5.6 mg) of(S)-3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amineobtained in Example 2 as an intermediate, 4-(dimethylamino)but-2-enoicacid hydrochloride (6.3 mg), and HATU (15 mg) were dissolved in DMF (1.0ml). DIPEA (50 μl) was added thereto, followed by stirring overnight.Chloroform and water were added to the reaction mixture to separate theorganic layer. After being washed with a saturated sodium chloridesolution, the organic layer was dried over anhydrous sodium sulfate, andthe solvent was then distilled off under reduced pressure. The resultingresidue was purified by preparative reversed-phase HPLC purification(water/acetonitrile (0.1% formic acid)) to obtain the title compound asa colorless, amorphous substance (2.3 mg). Table 1 shows the physicalproperties thereof.

Example 5 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-yn-1-one(Compound of Example 5)

The crude product (16 mg) of(S)-3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amineobtained in Example 2 as an intermediate, 3-(trimethylsilyl)propionicacid (10 mg), HATU (28 mg) were dissolved in DMF (0.5 ml). DIPEA (31 μl)was added thereto, followed by stirring overnight. Ethyl acetate and asaturated aqueous sodium bicarbonate solution were added to the reactionmixture to separate the organic layer. After drying the organic layerover anhydrous sodium sulfate, the solvent was distilled off underreduced pressure. The resulting residue was purified by silica gelcolumn chromatography (developing solvent: ethyl acetate/methanol) toobtain the title compound as a white solid (0.7 mg). Table 2 shows thephysical properties thereof.

Example 6 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)-2-fluoroprop-2-en-1-one(Compound of Example 6)

In accordance with Example 4, except that 2-fluoro-acrylic acid was usedin place of 4-(dimethylamino)but-2-enoic acid hydrochloride, the titlecompound was obtained as a colorless, amorphous substance. Table 2 showsthe physical properties thereof.

Example 7 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)-2-(pyrrolidin-1-ylmethyl)prop-2-en-1-one(Compound of Example 7)

In accordance with Example 4, except that2-(pyrrolidin-1-ylmethyl)acetic acid (Synth. Commun. 1995, 641) was usedin place of 4-(dimethylamino)but-2-enoic acid hydrochloride, the titlecompound was obtained as a colorless, amorphous substance. Table 2 showsthe physical properties thereof.

Example 8 Synthesis of(S)-1-(3-(4-amino-3-((3,5-diethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one(Compound of Example 8) (Step 1) Synthesis of1,3-diethoxy-5-ethynylbenzene

Carbon tetrabromide (4.78 g) was dissolved in dichloroethane (14 mL),and triphenylphosphine (7.56 g) was added thereto at 0° C. Afterstirring at 0° C. for 5 minutes, a solution of 3,5-diethoxybenzaldehyde(1.40 g) in dichloromethane (7 mL) was added thereto, followed bystirring for 20 minutes. Without performing further treatment, thereaction mixture was purified by silica gel chromatography (developingsolvent: hexane/ethyl acetate) to obtain1-(2,2-dibromovinyl)-3,5-diethoxybenzene. The obtained compound was usedfor the subsequent reaction without further purification. The compoundobtained above was dissolved in THF (30 mL). A 1.63 M n-butyllithiumsolution in hexane (10.5 mL) was added thereto at −78° C. The resultingmixture was stirred at −78° C. for 30 minutes. After adding a saturatedaqueous ammonium chloride solution, the reaction mixture was subjectedto extraction using ethyl acetate. The resulting organic layer waswashed with a saturated sodium chloride solution, and the organic layerwas then dried over anhydrous sodium sulfate. The solvent wasconcentrated under reduced pressure. The resulting residue was purifiedby silica gel column chromatography (developing solvent: hexane/ethylacetate) to obtain the title compound as a colorless, oily substance(1.31 g). Physical properties: m/z [M+H]⁺ 191.0

Step 2

In accordance with Example 1, except that 1,3-diethoxy-5-ethynylbenzeneobtained in Step 1 was used in place of 1-ethynyl-3,5-dimethoxybenzene,the title compound was obtained as a colorless, amorphous substance.Table 2 shows the physical properties thereof.

Example 9 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)prop-2-en-1-one(Compound of Example 9) (Step 1) Synthesis of tert-butyl3-(methylsulfonyloxy)azetidine-1-carboxylate

N-Boc-3-hydroxyazetidine (1.73 g) was dissolved in chloroform (20 ml).Triethylamine (2.09 ml) and methanesulfonyl chloride (856 μl) were addedthereto at 0° C. After stirring at room temperature for 0.5 hours, ethylacetate and water were added thereto to separate the organic layer.After being washed with a saturated aqueous sodium bicarbonate solution,a saturated aqueous ammonium chloride solution, and water, the organiclayer was dried over anhydrous sodium sulfate. The solvent was thendistilled off under reduced pressure to obtain the title compound as acolorless, oily compound (2.32 g). Physical properties: m/z [M+H]⁺ 252.0

(Step 2) Synthesis of tert-butyl3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidine-1-carboxylate

A suspension of tert-butyl 3-(methylsulfonyloxy)azetidine-1-carboxylate(1.32 g) obtained in Step 1 above,3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (1.37 g), and cesiumcarbonate (3.47 g) in DMF (10 ml) was stirred at 90° C. for 10 hours.Ethyl acetate and water were added thereto to separate the organiclayer. The organic layer was dried over anhydrous sodium sulfate, andthe solvent was distilled off under reduced pressure. The resultingresidue was purified by basic silica gel column chromatography(developing solvent: hexane/ethyl acetate) to obtain the title compoundas a light-yellow amorphous substance (482 mg). Physical properties: m/z[M+H]⁺ 417.0

(Step 3) Synthesis of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidine-1-carboxylate

PdCl₂ (dppf) CH₂Cl₂ (39 mg) was added to a mixture of tert-butyl3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidine-1-carboxylate(200 mg) obtained in Step 2 above, 1-ethynyl-3,5-dimethoxybenzene (117mg), copper (I) iodide (14 mg), and triethylamine (0.5 ml) in THF (5ml). After nitrogen purging, the resulting mixture was stirred at 80° C.for 1.5 hours. Ethyl acetate and water were added to the reactionmixture to separate the organic layer. After being washed with asaturated sodium chloride solution, the organic layer was dried overanhydrous sodium sulfate, and the solvent was distilled off underreduced pressure. The resulting residue was purified by basic silica gelcolumn chromatography (developing solvent: hexane/ethyl acetate) toobtain the title compound as a colorless, amorphous substance (185 mg).Physical properties: m/z [M+H]⁺ 451.1

(Step 4) Synthesis of Compound of Example 9

In accordance with Example 1 (Step 4), except that tert-butyl3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidine-1-carboxylateobtained in Step 3 above was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,a crude product of1-(azetidin-3-y)-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-aminewas obtained by removing a Boc group under acidic conditions.Thereafter, amidation was conducted to obtain the title compound as awhite solid. Table 2 shows the physical properties thereof.

Example 10 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-hydroxybut-2-yn-1-one(Compound of Example 10)

A crude product (6.0 mg) of1-(azetidin-3-yl)-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amineobtained in Example 9 as an intermediate, 4-hydroxybut-2-ynoic acid (3.7mg), and HATU (11 mg) were dissolved in DMF (1.0 ml). DIPEA (30 μl) wasadded thereto, followed by stirring overnight.

Chloroform and water were added to the reaction mixture to separate theorganic layer. After being washed with a saturated sodium chloridesolution, the organic layer was dried over anhydrous sodium sulfate, andthe solvent was distilled off under reduced pressure. The resultingresidue was purified by preparative reversed-phase HPLC purification(water/acetonitrile (0.1% formic acid)) to obtain the title compound asa colorless, amorphous substance (1.4 mg). Table 3 shows the physicalproperties thereof.

Example 11 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-hydroxy-4-methylpent-2-yn-1-one(Compound of Example 11)

In accordance with Example 10, except that4-hydroxy-4-methylpent-2-ynoic acid was used in place of4-hydroxybut-2-ynoic acid, the title compound was obtained as acolorless, amorphous substance. Table 3 shows the physical propertiesthereof.

Example 12 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Compound of Example 12)

In accordance with Example 10, except that 4-(dimethylamino)but-2-enoicacid hydrochloride was used in place of 4-hydroxybut-2-ynoic acid, thetitle compound was obtained as a colorless, amorphous substance. Table 3shows the physical properties thereof.

Example 13 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(cyclopropylamino)but-2-en-1-one(Compound of Example 13) (Step 1) Synthesis of 4-bromobut-2-enoylchloride

Thionyl chloride (3.0 ml) was added to 4-bromocrotonic acid (329 mg),and the mixture was stirred at 80° C. for 5 hours. The reaction mixturewas concentrated under reduced pressure, and toluene azeotropicdistillation was subsequently performed to obtain the title compound asa crude product (394 mg).

(Step 2) Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-bromobut-2-en-1-one

A crude product (140 mg) of1-(azetidin-3-yl)-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amineobtained in Example 9 as an intermediate was suspended in THF (4.5 ml),DIPEA (178 μl) was added thereto, and the mixture was cooled to 0° C. Asolution of 4-bromobut-2-enoyl chloride (66 mg) obtained in Step 1 abovein THF (0.5 ml) was added to the mixture dropwise and stirred at roomtemperature for 15 minutes. After halting the reaction using a saturatedaqueous sodium bicarbonate solution, the resulting product was extractedwith ethyl acetate. After drying the result over anhydrous sodiumsulfate, the solvent was distilled off under reduced pressure to obtainthe title compound as a crude product (160 mg). Physical properties: m/z[M+H]⁺ 497.0, 499.0

(Step 3) Synthesis of Compound of Example 13

The crude product (12 mg) of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-bromobut-2-en-1-oneobtained in Step 2 above was dissolved in DMF (0.5 ml). Cyclopropylamine(5 μl) and DIPEA (10 μl) were added thereto, and the mixture was stirredat room temperature for 1 hour. After concentrating under reducedpressure, the resulting residue was purified by preparativereversed-phase HPLC purification (water/acetonitrile (0.1% formic acid))to obtain the title compound as a colorless, amorphous substance (3.4mg). Table 3 shows the physical properties thereof.

Example 14 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(isopropylamino)but-2-en-1-one(Compound of Example 14)

In accordance with Example 13 (Step 3), except that isopropylamine wasused in place of cyclopropylamine, the title compound was obtained as acolorless, amorphous substance. Table 4 shows the physical propertiesthereof.

Example 15 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(ethyl(methyl)amino)but-2-en-1-one(Compound of Example 15)

In accordance with Example 13 (Step 3), except that ethylmethylamine wasused in place of cyclopropylamine, the title compound was obtained as acolorless, amorphous substance. Table 4 shows the physical propertiesthereof.

Example 16 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(cyclobutylamino)but-2-en-1-one(Compound of Example 16)

In accordance with Example 13 (Step 3), except that cyclobutylamine wasused in place of cyclopropylamine, the title compound was obtained as acolorless, amorphous substance. Table 4 shows the physical propertiesthereof.

Example 17 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(diethylamino)but-2-en-1-one(Compound of Example 17)

In accordance with Example 13 (Step 3), except that diethylamine wasused in place of cyclopropylamine, the title compound was obtained as acolorless, amorphous substance. Table 5 shows the physical propertiesthereof.

Example 18 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(tert-butylamino)but-2-en-1-one(Compound of Example 18)

In accordance with Example 13 (Step 3), except that tert-butylamine wasused in place of cyclopropylamine, the title compound was obtained as acolorless, amorphous substance. Table 5 shows the physical propertiesthereof.

Example 19 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(isopropyl(methyl)amino)but-2-en-1-one(Compound of Example 19)

In accordance with Example 13 (Step 3), except that isopropylmethylaminewas used in place of cyclopropylamine, the title compound was obtainedas a colorless, amorphous substance. Table 5 shows the physicalproperties thereof.

Example 20 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(piperidin-1-yl)but-2-en-1-one(Compound of Example 20)

In accordance with Example 13 (Step 3), except that piperidine was usedin place of cyclopropylamine, the title compound was obtained as acolorless, amorphous substance. Table 5 shows the physical propertiesthereof.

Example 21 Synthesis of1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-morpholinobut-2-en-1-one(Compound of Example 21)

In accordance with Example 13 (Step 3), except that morpholine was usedin place of cyclopropylamine, the title compound was obtained as acolorless, amorphous substance. Table 6 shows the physical propertiesthereof.

Example 22 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(3-fluoropyrrolidin-1-yl)but-2-en-1-one

In accordance with Example 13 (Step 3), except that(S)-3-fluoropyrrolidine was used in place of cyclopropylamine, the titlecompound was obtained as a colorless, amorphous substance. Table 6 showsthe physical properties thereof.

Example 23 Synthesis of(R)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(3-fluoropyrrolidin-1-yl)but-2-en-1-one

In accordance with Example 13 (Step 3), except that(R)-3-fluoropyrrolidine was used in place of cyclopropylamine, the titlecompound was obtained as a colorless, amorphous substance. Table 6 showsthe physical properties thereof.

Example 24 Synthesis of1-(3-(4-amino-3-((3,5-diethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)prop-2-en-1-one(Compound of Example 24)

In accordance with Example 9, except that 1,3-diethoxy-5-ethynylbenzenewas used in place of 1-ethynyl-3,5-dimethoxybenzene, the title compoundwas obtained as a colorless, amorphous substance. Table 7 shows thephysical properties thereof.

Example 25 Synthesis of1-(3-(4-amino-3-((3,5-diethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)azetidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Compound of Example 25)

In accordance with Example 12, except that 1,3-diethoxy-5-ethynylbenzenewas used in place of 1-ethynyl-3,5-dimethoxybenzene, the title compoundwas obtained as a colorless, amorphous substance. Table 7 shows thephysical properties thereof.

Example 26 Synthesis of1-(3-((4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)pyrrolidin-1-yl)prop-2-en-1-one(Compound of Example 26) (Step 1) Synthesis of tert-butyl3-((4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)pyrrolidine-1-carboxylate

DIAD (197 μl) was added to a suspension of3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine(148 mg) obtained in Example 1 (Step 1),N-Boc-3-hydroxymethylpyrrolidine (154 mg), polymer supportedtriphenylphosphine (upto 3.0 mmol/g, 334 mg) in THF (5.0 ml), followedby stirring at room temperature for 3 hours. The insoluble matter wasfiltered out, and the solvent was distilled off under reduced pressure.The resulting residue was purified by basic silica gel columnchromatography (developing solvent: hexane/ethyl acetate) to obtain thetitle compound as a colorless, amorphous substance (94.5 mg). Physicalproperties: m/z [M+H]⁺ 479.1

(Step 2) Synthesis of Compound of Example 26

In accordance with Example 1 (Step 4), except that tert-butyl3-((4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)pyrrolidine-1-carboxylateobtained in Step 1 above was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,the title compound was obtained as a white solid. Table 7 shows thephysical properties thereof.

Example 27 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one(Compound of Example 27)

In accordance with Example 26, except that (R)-1-Boc-3-hydroxypiperidinewas used in place of N-Boc-3-hydroxymethylpyrrolidine, the titlecompound was obtained as a white solid. Table 7 shows the physicalproperties thereof.

Example 28 Synthesis of1-((2S,4S)-4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-(hydroxymethyl)pyrrolidin-1-yl)prop-2-en-1-one(Compound of Example 28) (Step 1) Synthesis of (2S,4R)-tert-butyl2-((tert-butyldiphenylsilyloxy)methyl)-4-hydroxypyrrolidine-1-carboxylate

1-N-Boc-(2S,4R)-4-hydroxy-2-(hydroxymethyl)-pyrrolidine (500 mg) wasdissolved in DMF (4.0 ml), and imidazole (164 mg) was added to thedisolution. After cooling to 0° C., tert-butylchlorodiphenylsilane (616μl) was added to the mixture, and stirred for 1 hour. Ethyl acetate anda saturated aqueous sodium bicarbonate solution were added to thereaction mixture to separate the organic layer. After being washed witha saturated sodium chloride solution, the organic layer was dried overanhydrous sodium sulfate, and the solvent was distilled off underreduced pressure. The resulting residue was purified by silica gelcolumn chromatography (developing solvent: hexane/ethyl acetate) toobtain the title compound as a colorless, oily substance (655 mg).Physical properties: m/z [M+H]⁺ 456.2

(Step 2) Synthesis of (2S,4S)-tert-butyl2-((tert-butyldiphenylsilyloxy)methyl)-4-(methylsulfonyloxy)pyrrolidine-1-carboxylate

(2S,4R)-tert-Butyl2-((tert-butyldiphenylsilyloxy)methyl)-4-hydroxypyrrolidine-1-carboxylate(300 mg) obtained in Step 1 above was dissolved in chloroform (3.0 ml).Triethylamine (137 μl) and methanesulfonyl chloride (56 μl) were addedto the solution at 0° C. After stirring at room temperature for 2.0hours, chloroform and water were added thereto to separate the organiclayer. The organic layer was dried over anhydrous magnesium sulfate, andthe solvent was distilled off under reduced pressure to obtain the titlecompound as a colorless, oily compound (389 mg). Physical properties:m/z [M+H]⁺ 534.1

(Step 3) Synthesis of (2S,4S)-tert-butyl4-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-((tert-butyldiphenylsilyloxy)methyl)pyrrolidine-1-carboxylate

(2S,4S)-tert-butyl2-((tert-butyldiphenylsilyloxy)methyl)-4-(methylsulfonyloxy)pyrrolidine-1-carboxylate(389 mg) obtained in Step 2 above,3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (188 mg), and potassiumcarbonate (363 mg) were suspended in DMF (4.0 ml), followed by stirringovernight at 80° C. Ethyl acetate and water were added thereto toseparate the organic layer. The organic layer was dried over anhydrousmagnesium sulfate, and the solvent was distilled off under reducedpressure. The resulting residue was purified by basic silica gel columnchromatography (developing solvent: hexane/ethyl acetate) to obtain thetitle compound as a colorless, oily substance (191 mg). Physicalproperties: m/z [M+H]⁺ 699.1

(Step 4) Synthesis of (2S,4S)-tert-butyl4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-((tert-butyldiphenylsilyloxy)methyl)pyrrolidine-1-carboxylate

PdCl₂ (dppf) CH₂Cl₂ (13 mg) was added to a mixture of (2S,4S)-tert-butyl4-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-((tert-butyldiphenylsilyloxy)methyl)pyrrolidine-1-carboxylate(113 mg) obtained in Step 3 above, 1-ethynyl-3,5-dimethoxybenzene (52mg), copper (I) iodide (6 mg), and triethylamine (0.4 ml) in THF (4 ml).After nitrogen purging, the mixture was stirred at 85° C. for 3 hours.Ethyl acetate and water were added to the reaction mixture to separatethe organic layer. After washing with a saturated sodium chloridesolution, the organic layer was dried over anhydrous sodium sulfate, andthe solvent was distilled off under reduced pressure. The resultingresidue was purified by basic silica gel column chromatography(developing solvent: hexane/ethyl acetate) to obtain the title compoundas a colorless, amorphous substance (100 mg). Physical properties: m/z[M+H]⁺ 733.3

(Step 5) Synthesis of (2S,4S)-tert-butyl4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-(hydroxymethyl)pyrrolidine-1-carboxylate

(2S,4S)-tert-butyl4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-((tert-butyldiphenylsilyloxy)methyl)pyrrolidine-1-carboxylate(25 mg) obtained in Step 4 above was dissolved in THF (1.0 ml). Silicagel-supported tetrabutylammonium fluolide (upto 1.5 mmol/g, 34 mg) wasadded thereto, followed by stirring overnight. Silica gel-supportedtetrabutylammonium fluolide (upto 1.5 mmol/g, 30 mg) was further addedthereto, and the mixture was further stirred for 2 days. After filteringthe reagent off, the solvent was distilled off under reduced pressure.The resulting residue was purified by basic silica gel columnchromatography (developing solvent: hexane/ethyl acetate) to obtain thetitle compound as a colorless, amorphous substance (62 mg). Physicalproperties: m/z [M+H]⁺ 495.1

(Step 6) Synthesis of Compound of Example 28

In accordance with Example 1 (Step 4), except that4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-(hydroxymethyl)pyrrolidine-1-carboxylateobtained in Step 5 above was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,the title compound was obtained as a white solid. Table 8 shows thephysical properties thereof.

Example 29 Synthesis of1-(3-((4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)azetidin-1-yl)prop-2-en-1-one(Compound of Example 29)

In accordance with Example 26, except that1-Boc-3-hydroxymethylazetidine was used in place ofN-Boc-3-hydroxymethylpyrrolidine, the title compound was obtained as alight-yellow, amorphous substance. Table 8 shows the physical propertiesthereof.

Example 30 Synthesis ofN-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)cyclobutyl)acrylamide(Compound of Example 30)

In accordance with Example 1, except that tert-butyl3-hydroxycyclobutylcarbamate was used in place ofN-Boc-3-hydroxypyrrolidine, the title compound was obtained as alight-yellow, amorphous substance. Table 8 shows the physical propertiesthereof.

Example 31 Synthesis of1-(4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one(Compound of Example 31)

In accordance with Example 1, except that tert-butyl4-bromopiperidin-1-carboxylate was used in place of tert-butyl3-(methylsulfonyloxy)pyrrolidine-1-carboxylate, the title compound wasobtained as a white solid. Table 8 shows the physical propertiesthereof.

Example 32 Synthesis of1-((2S,4S)-4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynylpyrrolidin-1-yl)prop-2-en-1-one(Compound of Example 32) (Step 1) Synthesis of (2S,4S)-tert-butyl4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynylpyrrolidine-1-carboxylate

3-((3,5-Dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (14mg) obtained in Example 1 (Step 1), (2S,4R)-tert-butyl2-ethynyl-4-hydroxypyrrolidine-1-carboxylate (15 mg) synthesized by themethod disclosed in WO2005/007083, and triphenylphosphine (23 mg) weresuspended in THF (1.0 ml). DIAD (18 μl) was added to the suspension, andthe mixture was stirred at room temperature for 1 hour. The reactionmixture was concentrated and dissolved in a solution of DMSO. Theresulting solution was purified by preparative reversed-phase HPLCpurification (water/acetonitrile (0.1% formic acid)) to obtain the titlecompound as a colorless, amorphous substance (5.0 mg). Physicalproperties: m/z [M+H]⁺ 489.2

(Step 2) Synthesis of Compound of Example 32

In accordance with Example 1 (Step 4), except that (2S,4S)-tert-butyl4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)-2-ethynylpyrrolidine-1-carboxylateobtained in Step 1 above was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,the title compound was obtained as a white solid. Table 9 shows thephysical properties thereof.

Example 33 Synthesis of1-(4-((4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)piperidin-1-yl)prop-2-en-1-one(Compound of Example 33)

In accordance with Example 26, except that1-Boc-4-hydroxymethylpiperidine was used in place ofN-Boc-3-hydroxymethylpyrrolidine, the title compound was obtained as alight-yellow, amorphous substance. Table 9 shows the physical propertiesthereof.

Example 34 Synthesis ofN-(3-((4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)phenyl)acrylamide(Compound of Example 34)

In accordance with Example 26, except that (3-aminophenyl)methanol wasused in place of N-Boc-3-hydroxymethylpyrrolidine, the title compoundwas obtained as a colorless, amorphous substance. Table 9 shows thephysical properties thereof.

Example 35 Synthesis of1-(3-((4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)azetidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Compound of Example 35)

In accordance with Example 4, except that1-(azetidin-3-ylmethyl)-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine(i.e., the intermediate obtained in Example 29) was used in place of(S)-3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine,the title compound was obtained as a light-yellow, amorphous substance.Table 9 shows the physical properties thereof.

Example 36 Synthesis of1-(4-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Compound of Example 36)

In accordance with Example 4, except that3-((3,5-dimethoxyphenyl)ethynyl)-1-(piperidin-4-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine(i.e., the intermediate obtained in Example 31) was used in place of(S)-3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine,the title compound was obtained as a light-yellow, amorphous substance.Table 9 shows the physical properties thereof.

Example 37 Synthesis of1-(4-((4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)methyl)piperidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Compound of Example 37)

In accordance with Example 4, except that3-((3,5-dimethoxyphenyl)ethynyl)-1-(piperidin-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine(i.e., the intermediate obtained in Example 33) was used in place of(S)-3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine,the title compound was obtained as a light-yellow, amorphous substance.Table 10 shows the physical properties thereof.

Example 38 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Compound of Example 38) (Step 1) Synthesis of (S)-tert-butyl3-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylate

DIAD (1.41 ml) was added to a solution of4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (1.00 g),(R)—N-Boc-3-pyrrolidinol (1.01 g), and triphenylphosphine (1.88 g) intetrahydrofuran (40 ml), and the reaction mixture was stirred for 1hour. The reaction mixture was concentrated and washed with ethylacetate to obtain the title compound as a white solid (1.04 g). Physicalproperties: m/z [M+H]⁺ 448.9

(Step 2) Synthesis of (S)-tert-butyl3-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylate

Tetrahydrofuran (2.5 ml) and 28% aqueous ammonia (2.5 ml) were added to(S)-tert-butyl3-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylate(400 mg) obtained in Step 1 above. The reaction mixture was stirred at100° C. for 1.5 hours using a microwave reactor. Chloroform and waterwere added thereto to separate the organic layer. The organic layer wasdried over anhydrous sodium sulfate, and the solvent was distilled offunder reduced pressure to obtain the title compound as a white, solidcompound (382 mg). Physical properties: m/z [M+H]⁺ 430.3

(Step 3) Synthesis of (S)-tert-butyl3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylate

PdCl₂(dppf)₂CH₂Cl₂ (122 mg) was added to a mixture of (S)-tert-butyl3-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylate(660 mg) obtained in Step 2 above, 1-ethynyl-3,5-dimethoxybenzene (374mg), copper (I) iodide (44 mg), and triethylamine (2.0 ml) in THF (15ml). After nitrogen purging, the resulting mixture was stirred at 80° C.for 3.5 hours. Ethyl acetate and water were added to the reactionmixture to separate the organic layer. After washing with a saturatedsodium chloride solution, the organic layer was dried over anhydroussodium sulfate, and the solvent was distilled off under reducedpressure. The resulting residue was purified by basic silica gel columnchromatography (developing solvent: hexane/ethyl acetate) to obtain thetitle compound as a colorless, amorphous substance (714 mg). Physicalproperties: m/z [M+H]⁺ 464.1

(Step 4) Synthesis of Compound of Example 38

4N-Hydrochloric acid/1,4-dioxane (2 ml) was added to the (S)-tert-butyl3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylate(30 mg) obtained in Step 3 above, and the mixture was stirred at roomtemperature for 1.5 hours. After distilling the solvent of the resultingreaction mixture off under reduced pressure, toluene azeotropicdistillation was subsequently performed to obtain a crude product of(S)-5-((3,5-dimethoxyphenyl)ethynyl)-7-(pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(30 mg). A portion of the resulting crude product (10 mg),4-(dimethylamino)but-2-enoic acid hydrochloride (5.9 mg), and HATU (14mg) were dissolved in DMF (1.0 ml). DIPEA (50 μl) was added thereto andstirred at room temperature for 5 minutes. Chloroform and water wereadded to the reaction mixture to separate the organic layer. After beingwashed with a saturated sodium chloride solution, the organic layer wasdried over anhydrous sodium sulfate, and the solvent was distilled offunder reduced pressure. The resulting residue was purified bypreparative reversed-phase HPLC purification (water/acetonitrile (0.1%formic acid)) to obtain the title compound as a colorless, amorphoussubstance (3.9 mg). Table 10 shows the physical properties thereof.

Example 39 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)prop-2-en-1-one(Compound of Example 39)

In accordance with Example 1 (Step 4), except that (S)-tert-butyl3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylateobtained in Example 38 (Step 3) was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,a crude product of(S)-5-((3,5-dimethoxyphenyl)ethynyl)-7-(pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-aminewas obtained by removing a Boc group under acidic conditions.Thereafter, amidation was conducted to obtain the title compound as awhite solid. Table 10 shows the physical properties thereof.

Example 40 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(pyrrolidin-1-yl)but-2-en-1-one(Compound of Example 40) (Step 1) Synthesis of4-(pyrrolidin-1-yl)but-2-enoic acid hydrochloride

Methyl 4-bromocrotonate (1.79 g) was dissolved in tetrahydrofuran (40ml), pyrrolidine (1.67 ml) was added thereto at 0° C., and the mixturewas stirred at room temperature for 1 hour. Diethyl ether and water wereadded to the reaction mixture to separate the organic layer. After beingwashed with a saturated sodium chloride solution, the organic layer wasdried over anhydrous sodium sulfate, and the solvent was distilled offunder reduced pressure. 3N Hydrochloric acid (40 ml) was added to theresulting product, and the mixture was heated under reflux at 100° C.for 1 hour. The reaction mixture was concentrated under reducedpressure, and then the resulting residue was washed with a mixed solventof isopropanol and ethyl acetate to obtain the title compound as a whitesolid (939 mg). Physical properties: m/z [M+H]⁺ 156.0

(Step 2) Synthesis of Compound of Example 40

In accordance with Example 4 (Step 1), except that(S)-5-((3,5-dimethoxyphenyl)ethynyl)-7-(pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(the intermediate obtained in Example 38 (Step 4)) and4-(pyrrolidin-1-yl)but-2-enoic acid hydrochloride obtained in Step 1above were used, the title compound was obtained as a colorless,amorphous substance. Table 10 shows the physical properties thereof.

Example 41 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(4-methylpiperazin-1-yl)but-2-en-1-one(Compound of Example 41) (Step 1) Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-bromobut-2-en-1-one

The crude product (100 mg) of(S)-5-((3,5-dimethoxyphenyl)ethynyl)-7-(pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(i.e., the intermediate obtained in Example 38 (Step 4)) was suspendedin chloroform (3.0 ml). DIPEA (117 μl) was added to the suspension, andthe mixture was cooled to 0° C. A solution of 4-bromobut-2-enoylchloride (46 mg) obtained in Example 16 (Step 1) in chloroform (0.3 ml)was added thereto dropwise, and the resulting mixture was stirred atroom temperature for 15 minutes. After halting the reaction using asaturated aqueous sodium bicarbonate solution, the resulting product wasextracted with ethyl acetate. The organic layer was dried over anhydroussodium sulfate, and the solvent was distilled off under reduced pressureto obtain the title compound as a crude product (140 mg). Physicalproperties: m/z [M+H]⁺ 509.9, 511.9

(Step 2) Synthesis of Compound of Example 41

The crude product (12 mg) of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-bromobut-2-en-1-oneobtained in Step 1 above was dissolved in DMF (0.5 ml).N-methylpiperazine (4 mg) and DIPEA (10 μl) were added thereto, followedby stirring at room temperature overnight. After concentrating underreduced pressure, the resulting residue was purified by preparativereversed-phase HPLC purification (water/acetonitrile (0.1% formic acid))to obtain the title compound as a colorless, amorphous substance (3.0mg). Table 11 shows the physical properties thereof.

Example 42 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(4-hydroxypiperidin-1-yl)but-2-en-1-one(Compound of Example 42)

In accordance with Example 41 (Step 2), except that 4-hydroxypiperidinewas used in place of N-methylpiperazine, the title compound was obtainedas a colorless, amorphous substance. Table 11 shows the physicalproperties thereof.

Example 43 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(4-fluoropiperidin-1-yl)but-2-en-1-one(Compound of Example 43)

In accordance with Example 41 (Step 2), except that 4-fluoropiperidinewas used in place of N-methylpiperazine, the title compound was obtainedas a colorless, amorphous substance. Table 11 shows the physicalproperties thereof.

Example 44 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(3,3-difluoropyrrolidin-1-yl)but-2-en-1-one(Compound of Example 44)

In accordance with Example 41 (Step 2), except that3,3-difluoropyrrolidine was used in place of N-methylpiperazine, thetitle compound was obtained as a colorless, amorphous substance. Table12 shows the physical properties thereof.

Example 45 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(4,4-difluoropiperidin-1-yl)but-2-en-1-one(Compound of Example 45)

In accordance with Example 41 (Step 2), except that4,4-difluoropiperidine was used in place of N-methylpiperazine, thetitle compound was obtained as a colorless, amorphous substance. Table12 shows the physical properties thereof.

Example 46 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)but-2-yn-1-one(Compound of Example 46)

In accordance with Example 4 (Step 1), except that(S)-5-((3,5-dimethoxyphenyl)ethynyl)-7-(pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(i.e., the intermediate obtained in Example 38 (Step 4)) and 2-butynoicacid were used, the title compound was obtained as a colorless,amorphous substance. Table 12 shows the physical properties thereof.

Example 47 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-hydroxy-4-methylpent-2-yn-1-one(Compound of Example 47)

In accordance with Example 4 (Step 1), except that(5)-5-((3,5-dimethoxyphenyl)ethynyl)-7-(pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(i.e., the intermediate obtained in Example 38 (Step 4)) and4-hydroxy-4-methylpent-2-ynoic acid were used, the title compound wasobtained as a colorless, amorphous substance. Table 12 shows thephysical properties thereof.

Example 48 Synthesis of1-((S)-3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-((R)-3-fluoropyrrolidin-1-yl)but-2-en-1-one(Compound of Example 48)

In accordance with Example 41 (Step 2), except that(R)-3-fluoropyrrolidine was used in place of N-methylpiperazine, thetitle compound was obtained as a colorless, amorphous substance. Table13 shows the physical properties thereof.

Example 49 Synthesis of1-((S)-3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-((S)-3-fluoropyrrolidin-1-yl)but-2-en-1-one(Compound of Example 49)

In accordance with Example 41 (Step 2), except that(S)-3-fluoropyrrolidine was used in place of N-methylpiperazine, thetitle compound was obtained as a colorless, amorphous substance. Table13 shows the physical properties thereof.

Example 50 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidin-1-yl)-4-(piperidin-1-yl)but-2-en-1-one(Compound of Example 50)

In accordance with Example 41 (Step 2), except that piperidine was usedin place of N-methylpiperazine, the title compound was obtained as acolorless, amorphous substance. Table 13 shows the physical propertiesthereof.

Example 51 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)prop-2-en-1-one(Compound of Example 51) (Step 1) Synthesis of tert-butyl3-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidine-1-carboxylate

DIAD (1.41 ml) was added to a solution of4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (1.00 g),N-Boc-3-hydroxyazetidine (930 mg), and triphenylphosphine (1.85 g) intetrahydrofuran (40 ml), and the reaction mixture was stirred for 1hour. After concentrating, the reaction mixture was washed with ethylacetate to obtain the title compound as a white solid (1.07 g). Physicalproperties: m/z [M+H]⁺ 435.0

(Step 2) Synthesis of tert-butyl3-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidine-1-carboxylate

Tetrahydrofuran (2.5 ml) and 28% aqueous ammonia (2.5 ml) were added tothe tert-butyl3-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidine-1-carboxylate(350 mg) obtained in Step 1 above. The reaction mixture was stirred at100° C. for 1.5 hours using a microwave reactor. Chloroform and waterwere added thereto to separate the organic layer. The organic layer wasdried over anhydrous sodium sulfate, and the solvent was distilled offunder reduced pressure to obtain the title compound as a white solid(340 mg). Physical properties: m/z [M+H]⁺ 416.0

(Step 3) Synthesis of tert-butyl3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidine-1-carboxylate

PdCl₂(dppf)CH₂Cl₂ (122 mg) was added to a mixture of tert-butyl3-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidine-1-carboxylate(639 mg) obtained in Step 2 above, 1-ethynyl-3,5-dimethoxybenzene (374mg), copper (I) iodide (44 mg), and triethylamine (2.0 ml) in THF (15ml). After nitrogen purging, the resulting mixture was stirred at 80° C.for 3.5 hours. Ethyl acetate and water were added to the reactionmixture to separate the organic layer. After being washed with asaturated sodium chloride solution, the organic layer was dried overanhydrous sodium sulfate, and the solvent was distilled off underreduced pressure. The resulting residue was purified by basic silica gelcolumn chromatography (developing solvent: hexane/ethyl acetate) toobtain the title compound as a colorless, amorphous substance (704 mg).Physical properties: m/z [M+H]⁺ 450.1

(Step 4) Synthesis of Compound of Example 51

In accordance with Example 1 (Step 4), except that tert-butyl3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidine-1-carboxylateobtained in Step 3 was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,a crude product of7-(azetidin-3-y)-5-((3,5-)dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-4-aminewas obtained by removing a Boc group under acidic conditions.Thereafter, amidation was conducted to obtain the title compound as awhite solid. Table 13 shows the physical properties thereof.

Example 52 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Example Compound 52)

In accordance with Example 4 (Step 1), except that the tert-butyl3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidine-1-carboxylateobtained in Example 51 (Step 3) was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,the title compound was obtained as a colorless, amorphous substance.Table 14 shows the physical properties thereof.

Example 53 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(pyrrolidin-1-yl)but-2-en-1-one(Example Compound 53)

In accordance with Example 4 (Step 1), the7-(azetidin-3-yl)-5-((3,5-)dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amineobtained in Example 51 (Step 4) as an intermediate, and the4-(pyrrolidin-1-yl)but-2-enoic acid hydrochloride obtained in Example 40(Step 1) were used to obtain the title compound as a colorless,amorphous substance. Table 14 shows the physical properties thereof.

Example 54 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)but-2-yn-1-one(Example Compound 54)

In accordance with Example 4 (Step 1), the7-(azetidin-3-yl)-5-((3,5-)dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amineobtained as an intermediate in Example 51 (Step 4) and 2-butynoic acidwere used to obtain the title compound as a colorless, amorphoussubstance. Table 14 shows the physical properties thereof.

Example 55 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(azetidin-1-yl)but-2-en-1-one(Example Compound 55) (Step 1) Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-bromobut-2-en-1-one

A crude product of7-(azetidin-3-yl)-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(160 mg) obtained in Example 52 (Step 4) as an intermediate wassuspended in THF (3.0 ml). DIPEA (202 μl) was added thereto, and themixture was cooled to 0° C. A solution of 4-bromobut-2-enoyl chloride(75 mg) obtained in Example 16 (Step 1) in THF (0.5 ml) was added to themixture dropwise, and stirred at room temperature for 15 minutes. Afterhalting the reaction using a saturated sodium bicarbonate solution, theresulting product was extracted with ethyl acetate. After drying theresult over anhydrous sodium sulfate, the solvent was distilled offunder reduced pressure to obtain the title compound as a crude product(204 mg). Physical properties: m/z [M+H]⁺ 496.0, 498.0

(Step 2) Synthesis of Example Compound 55

The crude product of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-bromobut-2-en-1-one(10 mg) obtained in Step 1 above was suspended in THF (0.5 ml).Azetidine (7 μl) was added thereto, and the resulting mixture wasstirred for 1 hour at room temperature. After concentrating underreduced pressure, the resulting residue was purified by reversed-phaseHPLC purification (water/acetonitrile (0.1% formic acid)) to obtain thetitle compound as a colorless, amorphous substance (1.6 mg). Table 14shows the physical properties thereof.

Example 56 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(ethyl(methyl)amino)but-2-en-1-one(Example Compound 56)

In accordance with Example 55 (Step 2), except that ethylmethylamine wasused in place of azetidine, the title compound was obtained as acolorless, amorphous substance. Table 15 shows the physical propertiesthereof.

Example 57 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(isopropylamino)but-2-en-1-one(Example Compound 57)

In accordance with Example 55 (Step 2), except that isopropylamine wasused in place of azetidine, the title compound was obtained as acolorless, amorphous substance. Table 15 shows the physical propertiesthereof.

Example 58 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(isopropyl(methyl)amino)but-2-en-1-one(Example Compound 58)

In accordance with Example 55 (Step 2), except that isopropylmethylaminewas used in place of azetidine, the title compound was obtained as acolorless, amorphous substance. Table 15 shows the physical propertiesthereof.

Example 59 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(diethylamino)but-2-en-1-one(Example Compound 59)

In accordance with Example 55 (Step 2), except that diethylamine wasused in place of azetidine, the title compound was obtained as acolorless, amorphous substance. Table 16 shows the physical propertiesthereof.

Example 60 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-((2-methoxyethyl)(methyl)amino)but-2-en-1-one(Example Compound 60)

In accordance with Example 55 (Step 2), except that2-methoxy-N-methylethanamine was used in place of azetidine, the titlecompound was obtained as a colorless, amorphous substance. Table 16shows the physical properties thereof.

Example 61 Synthesis of1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(4-hydroxypiperidin-1-yl)but-2-en-1-one(Example Compound 61)

In accordance with Example 55 (Step 2), except that 4-hydroxypiperidinwas used in place of azetidine, the title compound was obtained as acolorless, amorphous substance. Table 16 shows the physical propertiesthereof.

Example 62 Synthesis of(S)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(3-hydroxypyrrolidin-1-yl)but-2-en-1-one(Example Compound 62)

In accordance with Example 55 (Step 2), except that(S)-3-hydroxypyrrolidine was used in place of azetidine, the titlecompound was obtained as a colorless, amorphous substance. Table 17shows the physical properties thereof.

Example 63 Synthesis of(R)-1-(3-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)azetidin-1-yl)-4-(3-hydroxypyrrolidin-1-yl)but-2-en-1-one(Example Compound 63)

In accordance with Example 55 (Step 2), except that(R)-3-hydroxypyrrolidine was used in place of azetidine, the titlecompound was obtained as a colorless, amorphous substance. Table 17shows the physical properties thereof.

Example 64 Synthesis of1-(4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)piperidin-1-yl)prop-2-en-1-one(Example Compound 64)

In accordance with Example 39, except that N-Boc-4-piperidinol was usedin place of (R)—N-Boc-3-pyrrolidinol, the title compound was obtained asa light-yellow, amorphous substance. Table 17 shows the physicalproperties thereof.

Example 65 Synthesis of1-(4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)piperidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Example Compound 65)

In accordance with Example 38, except that N-Boc-4-piperidinol was usedin place of (R)—N-Boc-3-pyrrolidinol, the title compound was obtained asa light-yellow, amorphous substance. Table 17 shows the physicalproperties thereof.

Example 66 Synthesis of (2S,4S)-methyl1-acryloyl-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate(Example Compound 66) (Step 1) Synthesis of4-chloro-5-iodo-7-((2-(trimethylsilyl)ethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidine

4-Chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (7.01 g) synthesized by themethod disclosed in WO2005/042556 was dissolved in anhydrous THF (125ml). After cooling to 0° C., 60% sodium hydride (4.02 g) was added tothe result, and the resulting mixture was stirred for 20 minutes.Subsequently, SEMCl (13.3 ml) was added thereto, and the mixture wasstirred at room temperature overnight. After cooling to 0° C. again,water was added to the mixture, and the reaction was halted. Theresulting product was extracted with ethyl acetate, and the organiclayer was washed with a saturated sodium chloride solution. After dryingthe result over anhydrous sodium sulfate, the solvent was distilled offunder reduced pressure. The resulting residue was purified by silica gelcolumn chromatography (developing solvent: hexane/ethyl acetate) toobtain the title compound as a white solid (7.28 g). Physicalproperties: m/z [M+H]⁺ 410.0

(Step 2) Synthesis of5-iodo-7-((2-(trimethylsilyl)ethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine

The4-chloro-5-iodo-7-((2-(trimethylsilyl)ethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidine(200 mg) obtained in Step 1 above was dissolved in THF (2.0 ml). 28%Aqueous ammonia (2 ml) was added thereto, and the reaction mixture wasthen stirred at 105° C. for 1.5 hours using a microwave reactor. Theresulting product was extracted with ethyl acetate, and the organiclayer was washed with a saturated sodium chloride solution. After dryingthe result over anhydrous sodium sulfate, the solvent was distilled offunder reduced pressure to obtain the title compound as a white solid(192 mg). Physical properties: m/z [M+H]⁺ 391.0

(Step 3) Synthesis of5-((3,5-dimethoxyphenyl)ethynyl)-7-((2-(trimethylsilyl)ethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine

In accordance with Example 1 (Step 1), except that the5-iodo-7-((2-(trimethylsilyl)ethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amineobtained in Step 2 above was used in place of3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine, the title compound wasobtained as a colorless solid. Physical properties: m/z [M+H]⁺425.4

(Step 4) Synthesis of5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine

A solution of5-((3,5-dimethoxyphenyl)ethynyl)-7-((2-(trimethylsilyl)ethoxy)methyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine(4.27 g) obtained in Step 3 in methylene chloride (20 ml) was cooled to0° C., and TFA (10 ml) was added thereto. The reaction mixture wasstirred at room temperature for 5 hours, and the solvent was distilledoff under reduced pressure. THF (50 ml) was added to the residue, andthe mixture was cooled to 0° C. 4N aqueous sodium hydroxide (12.5 ml)was added thereto, and the mixture was stirred at room temperatureovernight. The result was extracted with ethyl acetate, and dried overanhydrous magnesium sulfate. Subsequently, the solvent was distilled offunder reduced pressure, and chloroform was added to the resultingresidue. The mixture was subjected to filtration to obtain the titlecompound as a white solid (2.60 g). Physical properties: m/z [M+H]⁺295.3

(Step 5) Synthesis of(2S,4R)-4-(methylsulfonyloxy)-pyrrolidine-1,2-dicarboxylic acid1-tert-butylester-2-methylester

In accordance with Example 1 (Step 2), except that(2S,4R)-4-hydroxy-pyrrolidine-1,2-dicarboxylic acid1-tert-butylester-2-methylester was used in place ofN-Boc-3-pyrrolidinol, the title compound was obtained as a colorless,amorphous substance.

(Step 6) Synthesis of (2S,4S)-1-tert-butyl 2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate

In accordance with Example 1 (Step 3), except that the5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amineobtained in Step 4, the(2S,4R)-4-(methylsulfonyloxy)-pyrrolidine-1,2-dicarboxylic acid1-tert-butylester-2-methylester obtained in Step 5, sodium hydride, andNMP were individually used in place of3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine,tert-butyl 3-(methylsulfonyloxy)pyrrolidine-1-carboxylate, potassiumcarbonate, and DMF, the title compound was obtained as a colorless,amorphous substance. Physical properties: m/z [M+H]⁺ 522.4

(Step 7) Synthesis of (2S,4S)-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate

A solution of (2S,4S)-1-tert-butyl 2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate(24 mg) obtained in Step 6 above in methylene chloride (2.0 ml) and TFA(2.0 ml) was stirred at room temperature for 30 minutes. The solvent wasdistilled off under reduced pressure, and the resulting residue waspurified by basic silica gel column chromatography (developing solvent:chloroform/methanol). The title compound was thus obtained as alight-yellow, amorphous substance (11.9 mg). Physical properties: m/z[M+H]⁺ 422.1

(Step 8) Synthesis of Example Compound 66

Methylene chloride (2.0 ml) and triethylamine (16 μl) were added to the(2S,4S)-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate(12 mg) obtained in Step 7 above. After cooling to 0° C., chloroform(100 μl) in which acryloyl chloride (5 μl) was dissolved was added tothe resulting mixture, and stirred at room temperature for 10 minutes.After halting the reaction using a saturated sodium bicarbonatesolution, the resulting product was extracted with ethyl acetate. Afterdrying the result over anhydrous sodium sulfate, the solvent wasdistilled off under reduced pressure. The resulting residue was purifiedby silica gel column chromatography (developing solvent: ethylacetate/methanol) to obtain the title compound as a colorless, amorphoussubstance (4.2 mg). Table 18 shows the physical properties thereof.

Example 67 Synthesis of1-((2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((dimethylamino)methyl)pyrrolidin-1-yl)prop-2-en-1-one(Example Compound 67) (Step 1) Synthesis of (2S,4S)-tert-butyl2-((tert-butyldiphenylsilyloxy)methyl)-4-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylate

A solution of triphenylphosphine (443 mg) in THF (25 ml) was cooled to0° C., and DIAD (340 μl) was added thereto dropwise. The reactionmixture was stirred at 0° C. for 1 hour.4-Chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidine (363 mg) and the(2S,4R)-tert-butyl2-((tert-butyldiphenylsilyloxy)methyl)-4-hydroxypyrrolidine-1-carboxylate(651.6 mg) obtained in Example 28 (Step 1) were added thereto, andstirred at room temperature overnight. Ethyl acetate and water wereadded to separate the organic layer. The organic layer was dried overanhydrous magnesium sulfate, and the solvent was distilled off underreduced pressure. The resulting residue was purified by silica gelcolumn chromatography (developing solvent: hexane/ethyl acetate) toobtain the title compound as a light-yellow, amorphous substance (400mg). m/z [M+H]⁺ 718.5

(Step 2) Synthesis of (2S,4S)-tert-butyl4-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((tert-butyldiphenylsilyloxy)methyl)pyrrolidine-1-carboxylate

THF (10 ml) and an 8N ammonia methanol solution (5 ml) were added to the(2S,4S)-tert-butyl2-((tert-butyldiphenylsilyloxy)methyl)-4-(4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1-carboxylate(400 mg) obtained in Step 1. The mixture was stirred at 120° C. for 2hours under microwave irradiation, and the solvent was distilled offunder reduced pressure. The resulting residue was purified by silica gelcolumn chromatography (developing solvent: chloroform/ethanol) to obtainthe title compound as a light-yellow solid (293 mg). Physicalproperties: m/z [M+H]⁺ 698.5

(Step 3) Synthesis of (2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((tert-butyldiphenylsilyloxy)methyl)pyrrolidine-1-carboxylate

BoC₂O (188 mg) and DMAP (7 mg) were added to a solution of(2S,4S)-tert-butyl4-(4-amino-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((tert-butyldiphenylsilyloxy)methyl)pyrrolidine-1-carboxylate(200 mg) obtained in Step 2 in THF (5 ml), and the resulting mixture wasstirred at room temperature overnight. Ethyl acetate and water wereadded to the reaction mixture to separate the organic layer. The organiclayer was dried over anhydrous magnesium sulfate, and the solvent wasdistilled off under reduced pressure. The resulting residue was purifiedby silica gel column chromatography (developing solvent: hexane/ethylacetate) to obtain the title compound as a light-yellow, amorphoussubstance (238 mg). m/z [M+H]⁺ 898.5

(Step 4) Synthesis of (2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(hydroxymethyl)pyrrolidine-1-carboxylate

Silica gel-carrying tetrabutyl ammonium fluoride (700 mg) (up to 1.5mmol/g) was added to a solution of (2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((tert-butyldiphenylsilyloxy)methyl)pyrrolidine-1-carboxylate(237.5 mg) obtained in Step 3 in THF (5 ml), and the resulting mixturewas stirred at room temperature overnight. Silica gel was separated byfiltration, and the solvent of the filtrate was distilled off underreduced pressure. The resulting residue was purified by silica gelcolumn chromatography (developing solvent: hexane/ethyl acetate) toobtain the title compound as a light-yellow, amorphous substance (185mg). Physical properties: m/z [M+H]⁺ 660.2

(Step 5) Synthesis of (2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-formylpyrrolidine-1-carboxylate

Dess-Martin periodinane (51 mg) was added to a solution of(2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(hydroxymethyl)pyrrolidine-1-carboxylate(66 mg) obtained in Step 4 in methylene chloride (2 ml), and theresulting mixture was stirred at room temperature for 1 hour.Dess-Martin periodinane (100 mg) was further added to the mixture, andstirred at room temperature for 1 hour. Dess-Martin periodinane (70 mg)was additionally added thereto, and the resulting mixture was stirred atroom temperature for 1 hour. Water was added to the reaction mixture toseparate the organic layer. The organic layer was washed with a sodiumhydrogen carbonate aqueous solution and a 10% sodium thiosulfate aqueoussolution, and dried over anhydrous sodium sulfate. The solvent wasdistilled off under reduced pressure to obtain the title compound as alight-yellow, amorphous substance (68 mg). Physical properties: m/z[M+H]⁺658.1

(Step 6) Synthesis of (2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((dimethylamino)methyl)pyrrolidine-1-carboxylate

A 1M dimethylamine THF solution (0.3 ml) and acetic acid (0.2 ml) wereadded to a solution of (2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-formylpyrrolidine-1-carboxylate(68 mg) obtained in Step 5 in methylene chloride (2 ml), and theresulting mixture was cooled to 0° C. Sodium triacetoxyborohydride (127mg) was added to the reaction mixture, and stirred at 0° C. for 2 hours.The reaction mixture was neutralized using a sodium hydrogen carbonateaqueous solution, and the organic layer was dried over anhydrous sodiumsulfate. The solvent was distilled off under reduced pressure to obtainthe title compound as a light-yellow, amorphous substance (31.9 mg).Physical properties: m/z [M+H]⁺ 687.2

(Step 7) Synthesis of (2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((dimethylamino)methyl)pyrrolidine-1-carboxylate

In accordance with Example 1 (Step 1), except that the(2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((dimethylamino)methyl)pyrrolidine-1-carboxylateobtained in Step 6 was used in place of3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine, the title compound wasobtained as a light-yellow, amorphous substance. Physical properties:m/z [M+H]⁺ 721.5

(Step 8) Synthesis of5-((3,5-dimethoxyphenyl)ethynyl)-7-((3S,5S)-5-((dimethylamino)methyl)pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine

In accordance with Example 66 (Step 7), except that the(2S,4S)-tert-butyl4-(4-(bis(tert-butoxycarbonyl)amino)-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((dimethylamino)methyl)pyrrolidine-1-carboxylateobtained in Step 7 was used in place of (2S,4S)-1-tert-butyl 2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate,the title compound was obtained as a light-yellow, amorphous substance.Physical properties: m/z [M+H]⁺ 421.1

(Step 9) Synthesis of Example Compound 67

In accordance with Example 66 (Step 8), except that the5-((3,5-dimethoxyphenyl)ethynyl)-7-((3S,5S)-5-((dimethylamino)methyl)pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amineobtained in Step 8 was used in place of (2S,4S)-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate,the title compound was obtained as a colorless, amorphous substance.Table 18 shows the physical properties thereof.

Example 68 Synthesis of1-((2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(1,3,4-oxadiazole-2-yl)pyrrolidin-1-yl)prop-2-en-1-one(Example Compound 68) (Step 1) Synthesis of(2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-1-(tert-butoxycarbonyl)pyrrolidine-2-carboxylicacid

The (2S,4S)-1-tert-butyl 2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate(393.8 mg) obtained in Example 66 (Step 6) was dissolved in methanol (6ml). After cooling to 0° C., 4N aqueous sodium hydroxide (3 ml) wasadded thereto. The reaction suspension was stirred at room temperaturefor 3 hours. 5N hydrochloric acid was added to the reaction mixture to apH of 5, and ethyl acetate and water were added to separate the organiclayer. The organic layer was dried over anhydrous magnesium sulfate, andthe solvent was distilled off under reduced pressure to obtain the titlecompound as a light-yellow solid (280 mg). Physical properties: m/z[M+H]⁺ 508.3

(Step 2) Synthesis of (2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(hydrazinecarbonyl)pyrrolidine-1-carboxylate

DIPEA (73 μl) and hydrazine monohydrate (46 μl) were added to a solutionof(2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-1-(tert-butoxycarbonyl)pyrrolidine-2-carboxylicacid (106 mg) obtained in Step 1 above and TBTU (100 mg) in DMF (2 ml),and the resulting mixture was stirred at room temperature for 10minutes. Ethyl acetate and water were added to the reaction mixture toseparate the organic layer, and the organic layer was dried overanhydrous magnesium sulfate. The solvent was distilled off under reducedpressure to obtain the title compound as a light-yellow, amorphoussubstance (93.6 mg). Physical properties: m/z [M+H]⁺522.4

(Step 3) Synthesis of (2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(1,3,4-oxadiazole-2-yl)pyrrolidine-1-carboxylate

Toluene (3 ml) and trimethyl orthoformate (79 μl) were added to the(2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(hydrazinecarbonyl)pyrrolidine-1-carboxylate (93.6 mg) obtained in Step 2, and theresulting mixture was stirred at 110° C. overnight. Acetic acid (400 μl)was added to the reaction mixture, and stirred at 110° C. for 6 hours.Ethyl acetate and water were added to the reaction mixture to separatethe organic layer. The organic layer was dried over anhydrous magnesiumsulfate, and the solvent was distilled off under reduced pressure. Theresulting residue was purified by silica gel column chromatography(developing solvent: chloroform/methanol) to obtain the title compoundas a light-yellow, amorphous substance (50 mg). Physical properties: m/z[M+H]⁺ 532.2

(Step 4) Synthesis of7-((3S,5S)-5-(1,3,4-oxadiazole-2-yl)pyrrolidin-3-yl)-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine

In accordance with Example 66 (Step 7), except that the(2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(1,3,4-oxadiazole-2-yl)pyrrolidine-1-carboxylate(50 mg) obtained in Step 3 was used in place of (2S,4S)-1-tert-butyl2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate,the title compound was obtained as a light-yellow, amorphous substance(30.3 mg). m/z [M+H]⁺432.0

(Step 5) Synthesis of Example Compound 68

In accordance with Example 66 (Step 8), except that7-((3S,5S)-5-(1,3,4-oxadiazole-2-yl)pyrrolidin-3-yl)-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-4-amineobtained in Step 4 was used in place of (2S,4S)-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate,the title compound was obtained as a light-yellow, amorphous substance.Table 18 shows the physical properties thereof.

Example 69 Synthesis of1-((2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(5-methyl-1,3,4-oxadiazole-2-yl)pyrrolidin-1-yl)prop-2-en-1-one(Example Compound 69) (Step 1) Synthesis of (2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(5-methyl-1,3,4-oxadiazole-2-yl)pyrrolidine-1-carboxylate

In accordance with Example 68 (Step 3), except that triethylorthoacetate was used in place of trimethyl orthoformate, the titlecompound was obtained as a light-yellow, amorphous substance. Physicalproperties: m/z [M+H]⁺ 546.5

(Step 2) Synthesis of5-((3,5-dimethoxyphenyl)ethynyl)-7-((3S,5S)-5-(5-methyl-1,3,4-oxadiazole-2-yl)pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine

In accordance with Example 66 (Step 7), except that the(2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(5-methyl-1,3,4-oxadiazole-2-yl)pyrrolidine-1-carboxylateobtained in Step 1 was used in place of (2S,4S)-1-tert-butyl 2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate,the title compound was obtained as a light-yellow, amorphous substance.Physical properties: m/z [M+H]⁺ 466.0

(Step 3) Synthesis of Example Compound 69

In accordance with Example 66 (Step 8), except that the5-((3,5-dimethoxyphenyl)ethynyl)-7-((3S,5S)-5-(5-methyl-1,3,4-oxadiazole-2-yl)pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amineobtained in Step 2 was used in place of (2S,4S)-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate,the title compound was obtained as a light-yellow, amorphous substance.Table 18 shows the physical properties thereof.

Example 70 Synthesis of1-((2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(5-((dimethylamino)methyl)-1,3,4-oxadiazole-2-yl)pyrrolidin-1-yl)prop-2-en-1-one(Example Compound 70) (Step 1) Synthesis of (2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(2-(2-(dimethylamino)acetyl)hydrazinecarbonyl)pyrrolidine-1-carboxylate

In accordance with Example 68 (Step 2), except that2-(dimethylamino)acetohydrazide was used in place of hydrazinemonohydrate, the title compound was obtained as a light-yellow,amorphous substance. Physical properties: m/z [M+H]⁺ 607.3

(Step 2) Synthesis of (2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(5-((dimethylamino)methyl)-1,3,4-oxadiazole-2-yl)pyrrolidine-1-carboxylate

DIPEA (105 μl) and tosyl chloride (56 mg) were added to a solution of(2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(2-(2-(dimethylamino)acetyl)hydrazinecarbonyl)pyrrolidine-1-carboxylate (100 mg) obtained in Step 1 inacetonitrile (3 ml), and the mixture was stirred at 40° C. for 1 hour.Ethyl acetate and water were added to the reaction mixture to separatethe organic layer. The organic layer was dried over anhydrous magnesiumsulfate, and the solvent was distilled off under reduced pressure. Theresulting residue was purified by silica gel column chromatography(developing solvent: chloroform/methanol) to obtain the title compoundas a light-yellow, amorphous substance (40.5 mg). Physical properties:m/z [M+H]⁺ 589.2

Synthesis of (Step 3)5-((3,5-dimethoxyphenyl)ethynyl)-7-((3S,5S)-5-(5-((dimethylamino)methyl)-1,3,4-oxadiazole-2-yl)pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amine

In accordance with Example 66 (Step 7), except that the(2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-(5-((dimethylamino)methyl)-1,3,4-oxadiazole-2-yl)pyrrolidine-1-carboxylateobtained in Step (2) was used in place of (2S,4S)-1-tert-butyl 2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate,the title compound was obtained as a light-yellow, amorphous substance.Physical properties: m/z [M+H]⁺ 489.2

(Step 4) Synthesis of Example Compound 70

In accordance with Example 66 (Step 8), except that the5-((3,5-dimethoxyphenyl)ethynyl)-7-((3S,5S)-5-(5-((dimethylamino)methyl)-1,3,4-oxadiazole-2-yl)pyrrolidin-3-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-amineobtained in Step 3 was used in place of (2S,4S)-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate,the title compound was obtained as a light-yellow, amorphous substance.Table 19 shows the physical properties thereof.

Example 71 Synthesis of(2S,4S)-1-acryloyl-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-N-(2-(dimethylamino)ethyl)-N-methylpyrrolidine-2-carboxamide(Example Compound 71) (Step 1) Synthesis of (2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((2-(dimethylamino)ethyl)(methyl)carbamoyl)pyrrolidine-1-carboxylate

A solution of(2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-1-(tert-butoxycarbonyl)pyrrolidine-2-carboxylicacid (25.4 mg) obtained in Example 68 (Step 1), TBTU (17.7 mg),N,N,N′-trimethylethane-1,2-diamine (13 μl), and DIPEA (26 μl) inacetonitrile (3 ml) was stirred at room temperature for 1 hour. Ethylacetate and water were added to separate the organic layer. The organiclayer was dried over anhydrous sodium sulfate, and the solvent wasdistilled off under reduced pressure. The resulting residue was purifiedby basic silica gel column chromatography (developing solvent:chloroform/methanol) to obtain the title compound as a light-yellow,amorphous substance (3 mg). Physical properties: m/z [M+H]⁺ 592.4

(Step 2) Synthesis of(2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-N-(2-(dimethylamino)ethyl)-N-methylpyrrolidine-2-carboxamide

In accordance with Example 66 (Step 7), except that the(2S,4S)-tert-butyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-2-((2-(dimethylamino)ethyl)(methyl)carbamoyl)pyrrolidine-1-carboxylateobtained in Step 1 above was used in place of (2S,4S)-1-tert-butyl2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate,the title compound was obtained as a colorless, amorphous substance.Physical properties: m/z [M+H]⁺ 492.4

(Step 3) Synthesis of Example Compound 71

In accordance with Example 66 (Step 8), except that the(2S,4S)-4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-N-(2-(dimethylamino)ethyl)-N-methylpyrrolidine-2-carboxamideobtained in Step 2 was used in place of (2S,4S)-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-2-carboxylate,the title compound was obtained as a colorless, amorphous substance.Table 19 shows the physical properties thereof.

Example 72 Synthesis of1-(4-((4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)methyl)piperidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Example Compound 72)

In accordance with Example 38, except thatN-Boc-4-hydroxymethylpiperidin was used in place of(R)—N-Boc-3-pyrrolidinol, the title compound was obtained as alight-yellow, amorphous substance. Table 19 shows the physicalproperties thereof.

Example 73 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one (Example Compound 73)(Step 1) Synthesis of 4-chloro-3-iodo-1H-pyrrolo[3,2-c]pyridine

4-Chloro-1H-pyrrolo[3,2-c]pyridine (247 mg) synthesized by the methoddisclosed in WO2007/095223 was dissolved in DMF (7.0 ml). After coolingto 0° C., N-iodosuccinimide (382 mg) was added thereto. The resultingmixture was stirred at room temperature for 1 hour, and then chloroformand water were added thereto to separate the organic layer. After theorganic layer was dried over anhydrous magnesium sulfate, the solventwas distilled off under reduced pressure. The resulting residue waspurified by silica gel column chromatography (developing solvent:hexane/ethyl acetate) to obtain the title compound as a dark-brown solid(455 mg). Physical properties: m/z [M+H]⁺ 279.1

(Step 2) Synthesis of (S)-tert-butyl3-(4-chloro-3-iodo-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidine-1-carboxylate

The 4-chloro-3-iodo-1H-pyrrolo[3,2-c]pyridine (225 mg) obtained in Step1 was dissolved in DMF (3.0 ml). After cooling to 0° C., 60% sodiumhydride (64.5 mg) was added thereto. The (R)-tert-butyl3-(methylsulfonyloxy)pyrrolidine-1-carboxylate (322 mg) obtained inExample 2 (Step 1) was added to the reaction mixture using DMF (2.0 ml),and the mixture was stirred overnight. 60% Sodium hydride (64.5 mg) wasadditionally added, and the mixture was stirred at 85° C. overnight.Ethyl acetate and water were added to the reaction mixture to separatethe organic layer. The organic layer was dried over anhydrous magnesiumsulfate, and the solvent was distilled off under reduced pressure. Theresulting residue was purified by silica gel column chromatography(developing solvent: hexane/ethyl acetate) to obtain the title compoundas a crude product (192 mg). Physical properties: m/z [M+H]⁺ 448.3

(Step 3) Synthesis of (S)-tert-butyl3-(4-chloro-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidine-1-carboxylate

PdCl₂(dppf)CH₂Cl₂ (33 mg) was added to a mixture of the crude product of(S)-tert-butyl3-(4-chloro-3-iodo-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidine-1-carboxylate(180 mg) obtained in Step 2, 1-ethynyl 3,5-dimethoxybenzene (97 mg),copper (I) iodide (15 mg), and triethylamine (1.0 ml) in THF (4.0 ml).After nitrogen purging, the resulting mixture was stirred at 50° C. for30 minutes. Ethyl acetate and water were added to the reaction mixtureto separate the organic layer. After being washed with a saturatedsodium chloride solution, the organic layer was dried over anhydroussodium sulfate, and the solvent was then distilled off under reducedpressure. The resulting residue was purified by silica gel columnchromatography (developing solvent: hexane/ethyl acetate) to obtain thetitle compound as a colorless, amorphous substance (133 mg). Physicalproperties: m/z [M+H]⁺ 482.4

(Step 4) Synthesis of (S)-tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidine-1-carboxylate

Under a nitrogen atmosphere, the (S)-tert-butyl3-(4-chloro-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidine-1-carboxylate(120 mg) obtained in Step 3,2,2′-bis(diphenylphosphino)-1,1′-binaphthalene (BINAP) (26 mg), sodiumtert-butoxide (72 mg), benzophenone imine (92 mg), and tris(dibenzylideneacetone)dipalladium (36 mg) were suspended in toluene (10ml), and the result was stirred at 115° C. for 90 minutes. Afterdilution with ethyl acetate, celite filtration was performed. Thesolvent was distilled off under reduced pressure.Hydroxyaminehydrochloride (366 mg), sodium bicarbonate (442 mg),methanol (16 ml), and water (4 ml) were added to the resulting residue,and the resulting mixture was stirred at room temperature for 2 hours.The solvent was distilled off under reduced pressure. Thereafter, ethylacetate and a saturated sodium chloride solution were added to separatethe organic layer. The organic layer was dried over anhydrous sodiumsulfate, and the solvent was distilled off under reduced pressure. Theresulting residue was purified by basic silica gel column chromatography(developing solvent: hexane/ethyl acetate) to obtain the title compoundas a colorless, amorphous substance (35 mg). Physical properties: m/z[M+H]⁺ 463.4

(Step 5) Synthesis of Example Compound 73

In accordance with Example 1 (Step 4), except that the (S)-tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrrolo[3,2-c]pyridin-1-yl)pyrrolidine-1-carboxylateobtained in Step (3) was used in place of tert-butyl3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate,the title compound was obtained as a white solid. Table 19 shows thephysical properties thereof.

Reference Example 1 Synthesis of(R)-1-(3-(4-amino-3-(4-phenoxyphenyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)piperidin-1-yl)prop-2-en-1-one(Reference Example Compound 1)

The compound was synthesized according to the method disclosed inWO2008/121742. Table 20 shows the physical properties thereof.

Reference Example 2 Synthesis of 3-cyclobutyl-1-(phenylethynyl)imidazo[1,5-a]pyrazin-8-amine (Reference Example Compound 2)

The compound was synthesized according to the method disclosed inWO2007/087395. Table 20 shows the physical properties thereof.

Reference Example 3 Synthesis of(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)propan-1-one(Reference Example Compound 3)

In accordance with Example 1,(S)-3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amineand propionyl chloride were used to obtain the title compound as a whitesolid. Table 20 shows the physical properties thereof.

Reference Example 4 Synthesis of(S)-1-(3-(4-amino-3-((3,5-diisopropylphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Reference Example Compound 4) (Step 1) Synthesis of1-ethynyl-3,5-diisopropylbenzene

PdCl₂(dppf)CH₂Cl₂ (163 mg) was added to a mixture oftrimethylsilylacetylene (589 mg), 1-bromo-3,5-diisopropylbenzene (480mg), copper (I) iodide (76 mg), and triethylamine (0.11 ml) in THF (4ml). After nitrogen purging, the resulting mixture was stirred at 80° C.for 4 hours. Ethyl acetate and water were added to the reaction mixtureto separate the organic layer. After drying the result over anhydroussodium sulfate, the solvent was distilled off under reduced pressure. A2% potassium hydroxide methanol solution (10 ml) was added to theresulting residue, and the result was stirred at room temperatureovernight. Chloroform and water were added to the reaction mixture toseparate the organic layer. After drying the result over anhydroussodium sulfate, the solvent was distilled off under reduced pressure.The resulting residue was purified by silica gel column chromatography(developing solvent: hexane) to obtain the title compound as a yellow,oily substance (181 mg).

(Step 2) Synthesis of (S)-tert-butyl3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate

A suspension of 3-iodo-1H-pyrazolo[3,4-d]pyrimidin-4-amine (446 mg),(R)-tert-butyl 3-(methylsulfonyloxy)pyrrolidine-1-carboxylate (450 mg),potassium carbonate (692 mg) in DMF (5.0 ml) was stirred at 85° C. for 6hours. Ethyl acetate and water were added thereto to separate theorganic layer. The organic layer was dried over anhydrous sodiumsulfate, and the solvent was distilled off under reduced pressure. Theresulting residue was purified by basic silica gel column chromatography(developing solvent: hexane/ethyl acetate) to obtain the title compoundas a light-yellow, amorphous substance (354 mg). Physical properties:m/z [M+H]⁺ 431.1

(Step 3) Synthesis of (S)-tert-butyl3-(4-amino-3-((3,5-diisopropylphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate

PdCl₂(dppf)CH₂Cl₂ (8.2 mg) was added to a mixture of1-ethynyl-3,5-diisopropylbenzene (56 mg) obtained in Step 1,(S)-tert-butyl3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate(43 mg) obtained in Step 2, copper (I) iodide (3.8 mg), andtriethylamine (0.2 ml) in THF (2.0 ml). After nitrogen purging, theresulting mixture was stirred at 80° C. for 1.5 hours. Ethyl acetate andwater were added to the reaction mixture to separate the organic layer.The organic layer was washed with a saturated sodium chloride solution.After drying the result over anhydrous sodium sulfate, the solvent wasdistilled off under reduced pressure. The resulting residue was purifiedby basic silica gel column chromatography (developing solvent:hexane/ethyl acetate) to obtain the title compound as a colorless,amorphous substance (42 mg). Physical properties: m/z [M+H]⁺ 489.2

(Step 4) Synthesis of(S)-3-((3,5-diisopropylphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine

In accordance with Example 66 (Step 7), except that the (S)-tert-butyl3-(4-amino-3-((3,5-diisopropylphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylateobtained in Step (3) was used in place of (2S,4S)-1-tert-butyl 2-methyl4-(4-amino-5-((3,5-dimethoxyphenyl)ethynyl)-7H-pyrrolo[2,3-d]pyrimidin-7-yl)pyrrolidine-1,2-dicarboxylate,the title compound was obtained as a light-yellow, amorphous substance.

(Step 5) Synthesis of Reference Example Compound 4

In accordance with Example 4, except that the(S)-3-((3,5-diisopropylphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amineobtained in Step (4) was used in place of(S)-3-((3,5-dimethoxyphenyl)ethynyl)-1-(pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine,the title compound was obtained as a light-yellow, amorphous substance.Table 20 shows the physical properties thereof.

Reference Example 5 Synthesis of(S)-1-(3-(4-amino-3-((3-methoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Reference Example Compound 5) (Step 1) Synthesis of(S)-1-(3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one

4N-Hydrochloric acid/1,4-dioxane (4 ml) was added to the (S)-tert-butyl3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidine-1-carboxylate(488 mg) obtained in Reference Example 4 (Step 2), and the resultingmixture was stirred for 1 hour. The solvent was distilled off underreduced pressure. A solution of 4-(dimethylamino)but-2-enoic-acidhydrochloride (281 mg) and HATU (647 mg) in DMF (5.0 ml) was added tothe resulting residue. Further, DIPEA (0.78 ml) was added thereto, andthe mixture was stirred overnight. The reaction mixture was concentratedunder reduced pressure, and chloroform (50 ml) and ethanol (50 ml) wereadded to the resulting residue. The insoluble matter was removed byfiltration, and the filtrate was concentrated under reduced pressure.The resulting residue was washed with ethyl acetate (5.0 ml) and driedto obtain the crude product of the title compound (458 mg). Physicalproperties: m/z [M+H]⁺ 442.0

(Step 2) Synthesis of Reference Example Compound 5

PdCl₂(dppf)CH₂Cl₂ (1.3 mg) was added to a mixture of(S)-1-(3-(4-amino-3-iodo-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one(8.0 mg) obtained in Step 1, 1-ethynyl-3-methoxybenzene (4.0 mg), copper(I) iodide (0.6 mg), and triethylamine (8.6 μl) in THF (1.0 ml). Afternitrogen purging, the resulting mixture was stirred at 80° C. overnight.The reaction mixture was diluted with ethyl acetate and methanol. Theresulting diluted solution was treated with basic silica gel, and thenconcentrated. The resulting residue was purified by reversed-phase HPLCpurification (water/acetonitrile (0.1% formic acid)) to obtain the titlecompound as a colorless, amorphous substance (1.4 mg). Table 20 showsthe physical properties thereof.

Reference Example 6 Synthesis of(S)—N-(3-((4-amino-1-(1-(4-(dimethylamino)but-2-enoyl)pyrrolidin-3-yl)-1H-pyrazolo[3,4-d]pyrimidin-3-yl)ethynyl)phenyl)acetamide(Reference Example Compound 6)

In accordance with Reference Example 5, except thatN-(3-ethynylphenyl)acetamide was used in place of1-ethynyl-3-methoxybenzene, the title compound was obtained as acolorless, amorphous substance. Table 20 shows the physical propertiesthereof.

Reference Example 7 Synthesis of(S)-1-(3-(4-amino-3-(pyridin-3-ylethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)-4-(dimethylamino)but-2-en-1-one(Reference Example Compound 7)

In accordance with Reference Example 5, 3-ethynyl pyridine was used inplace of 1-ethynyl-3-methoxybenzene, the title compound was obtained asa colorless, amorphous substance. Table 20 shows the physical propertiesthereof.

Table 1

TABLE 1 Ex. Comp Structural formula Physical properties 1

1H-NMR (DMSO-d6) δ: 2.29-2.50 (2H, m), 3.55- 4.10 (4H, m), 3.77(6H, s),5.40-5.55 (1H, m), 5.62-5.72 (1H, m), 6.10-6.20 (1H, m), 6.50-6.70 (1H,m), 6.64(1H, d, J = 2.3 Hz), 6.90 (2H, d, J = 2.3 Hz), 8.26 (1H, s).m/z[M + H]⁺ 419.0 2

1H-NMR (DMSO-d6) δ: 2.29-2.50 (2H, m), 3.55- 4.10 (4H, m), 3.77(6H, s),5.40-5.55 (1H, m), 5.62-5.72 (1H, m), 6.10-6.20 (1H, m), 6.50-6.70 (1H,m), 6.64(1H, d, J = 2.3 Hz), 6.90 (2H, d, J = 2.3 Hz), 8.26 (1H, s).m/z[M + H]⁺ 419.0 3

1H-NMR (DMSO-d6) δ: 1.81 (1.5H, dd, J = 6.8, 1.6 Hz), 1.85 (1.5H, dd, J= 6.8, 1.6 Hz), 2.29-2.50 (2H, m), 3.56-3.92 (3.5H, m), 3.77(6H, s),4.02-4.10(0.5H, m), 5.12-5.53 (1H, m), 6.26 (0.5H, dd, J = 15.1, 1.6Hz), 6.34 (0.5H, dd, J = 15.1, 1.6 Hz), 6.60 (1H, t, J = 2.4 Hz),6.64-6.74 (1H, m), 6.91 (2H, d, J = 2.4 Hz), 8.25 (6.5H, S), 8.27 (0.5H,S). m/z[M + H]⁺ 433.1 4

1H-NMR (DMSO-d6) δ: 2.06 (3H, s), 2.09 (3H, s), 2.20-2.45 (2H, m), 2.95(1H, d, J = 5.9 Hz), 2.99 (1H, d, J = 5.9 Hz), 3.30-4.10 (4H, m),5.30-5.50 (1H, m), 6.29 (0.5H, d, J = 15.0 Hz), 6.38 (0.5H, d, J = 15.0Hz) 6.53-6.65 (3H, m), 6.84 (2H, s), 8.12 (1H, s). m/z[M + H]⁺ 476.1

TABLE 2 5

m/z[M + H]⁺ 417.0 6

1H-NMR (CDCl3) δ: 2.37-2.68(2H, m), 3.74- 3.82(2H, m), 3.82(6H, s),4.07-4.21(2H, m), 5.11- 5.19(1H, m), 5.50-5.56(1H, m), 5.58(1H, t, J =46.8, 2.0 Hz), 5.85(2H, s), 6.54(1H, t, J = 2.0 Hz), 6.74(2H, d, J = 2.4Hz), 8.37(1H, s) m/z[M + H]⁺ 437.2 7

1H-NMR (CDCl3) δ: 1.70-1.80(4H, m), 2.42 2.66(6H, m), 3.20-3.32(1H, m),3.40-3.46(1H, m), 3.76-3.82(2H, m), 3.82(6H, s), 3.91-4.15(3H, m),5.32-5.56(3H, m), 5.91(2H, s), 6.54(1H, t, J = 2.0 Hz) 6.73(2H, d, J =2.4 Hz), 8.36 (1H, s) m/z[M + H]⁺ 502.2 8

1H-NMR (CDCl3) δ: 1.42 (6H, t, J = 6.8 Hz), 2.39-2.72 (2H, m), 3.71-3.84(1H, m), 3.96-4.12 (3H, m), 4.03 (4H, q, J = 6.8 Hz), 5.48-5.76 (2H, m),5.84 (2H, br s), 6.38-6.56 (3H, m), 6.70-6.73 (2H, m), 8.36-8.38 (1H,m). m/z[M + H]⁺ 447.2 9

1H-NMR (DMSO-d6) δ: 3.78 (6H, s), 4.20-4.35 (1H, m), 4.40-4.50 (1H, m),4.55-4.65 (1H, m), 4.70-4.80 (1H, m), 5.70-5.80 (2H, m), 6.16 (1H, dd, J= 17.1, 2.1 Hz), 6.38 (1H, dd, J = 17.1, 10.2 Hz), 6.61 (1H, t, J = 2.4Hz), 6.94 (2H, d, J = 2.4 Hz), 8.26 (1H, s). m/z[M + H]⁺ 405.1

TABLE 3 10

1H-NMR (DMSO-d6) δ: 3.73 (6H, s), 4.12-4.28 (3H, m), 4.35-4.50 (2H, m),4.55-4.65 (1H, m), 5.55(1H, brs), 5.67 (1H, brs), 6.56 (1H, s), 6.87(2H, s), 8.40 (1H, s). m/z[M + H]⁺ 433.3 11

1H-NMR (DMSO-d6) δ: 1.32 (6H, s), 3.70 (6H, s), 4.14-4.19 (1H, m),4.33-4.44 (2H, m), 4.57 (1H, t, J = 9.3 Hz), 5.61-5.70 (2H, m), 6.53(1H, s), 6.87 (2H, d, J = 2.2 Hz), 8.18 (1H, s). m/z[M + H]⁺ 461.4 12

1H-NMR (DMSO-d6) δ: 2.08 (6H, s), 2.97 (2H, d, J = 6.2 Hz), 3.72 (6H, d,J = 3.7 Hz), 4.15- 4.25 (1H, m), 4.30-4.48 (2H, m), 4.67 (1H, t, J = 9.0Hz), 5.60-5.70 (1H, m), 6.11 (1H, d, J = 15.0 Hz), 6.55-6.61 (2H, m),6.88 (2H, d, J = 1.8 Hz), 8.20 (1H, s). m/z[M + H]⁺ 462.1 13

1H-NMR (DMSO-d6) δ: 0.00-0.03 (2H, m), 0.12- 0.18 (2H, m), 1.84-1.90(1H, m), 3.59 (6H, s), 4.04-4.09 (1H, m), 4.22 (1H, t, J = 9.3 Hz),4.32-4.38 (1H, m), 4.52 (1H, t, J = 8.4 Hz), 5.48-5.54 (1H, m), 5.93(1H, d, J = 15.4 Hz), 6.41 (1H, t, J = 2.3 Hz), 6.54 (1H, dt, J = 15.4,5.5 Hz), 6.75 (2H, d, J = 2.3 Hz), 8.07 (1H, s). m/z[M + H]⁺ 474.2

TABLE 4 14

1H-NMR (DMSO-d6) δ: 0.98 (6H, d, J = 6.2 Hz), 2.70-2.76 (1H, m), 3.79(6H, s), 4.24-4.29 (1H, m), 4.43 (1H, t, J = 9.3 Hz), 4.54-4.58 (1H, m),4.72 (1H, t, J = 8.6 Hz), 5.70-5.74 (1H, m), 6.17 (1H, d, J = 15.4 Hz),6.61 (1H, t, J = 2.5 Hz), 6.68-6.80 (1H, m), 6.95 (2H, d, J = 2.5 Hz),8.33 (1H, s). m/z[M + H]⁺ 476.2 15

1H-NMR (DMSO-d6) δ: 0.98 (3H, t, J = 7.1 Hz), 2.13 (3H, s), 2.36 (2H, q,J = 7.1 Hz), 3.11 (2H, d, J = 6.2 Hz), 3.79 (6H, s), 4.23- 4.30(1H, m),4.43 (1H, t, J = 9.2 Hz), 4.50- 4.58 (1H, m), 4.73 (1H, t, J = 8.8 Hz),5.70- 5.74 (1H, m), 6.18 (1H, d, J = 15.4 Hz), 6.60- 6.70 (2H, m), 6.95(2H, d, J = 2.6 Hz), 8.28 (1H, s). m/z[M + H]⁺ 476.2 16

1H-NMR (DMSO-d6) δ: 1.50-1.70 (4H, m), 2.02- 2.10 (2H, m), 3.08-3.15(1H, m), 3.20-3.25 (2H, m), 3.79 (6H, s), 4.24-4.29 (1H, m), 4.42 (1H,t, J = 9.0 Hz), 4.53-4.58 (1H, m), 4.72 (1H, t, J = 8.6 Hz), 5.70-5.74(1H, m), 6.13 (1H, d, J = 15.4 Hz), 6.61 (1H, t, J = 2.3 Hz), 6.67-6.74(1H, m), 6.95 (2H, d, J = 2.3 Hz), 8.27 (1H, s). m/z[M + H]⁺ 488.1

TABLE 5 17

1H-NMR (DMSO-d6) δ: 0.96 (6H, t, J = 7.1 Hz), 2.42-2.49 (4H, m), 3.19(2H, d, J = 4.8 Hz), 3.79 (6H, s), 4.25-4.30 (1H, m), 4.43 (1H, t, J =9.2 Hz), 4.52-4.57 (1H, m), 4.73 (1H, t, J = 8.6 Hz), 5.69-5.74 (1H, m),6.19 (1H, d, J = 15.4 Hz), 6.60-6.72 (2H, m), 6.95 (2H, d, J = 2.2 Hz),8.26 (1H, s). m/z[M + H]⁺ 490.2 18

1H-NMR (DMSO-d6) δ: 1.07 (9H, s), 3.78 (6H, s), 4.24-4.29 (1H, m), 4.43(1H, t, J = 9.5 Hz), 4.50-4.60 (1H, m), 4.72 (1H, t, J = 8.2 Hz),5.68-5.75 (1H, m), 6.21 (1H, d, J = 15.4 Hz), 6.61 (1H, t, J = 2.3 Hz),6.70 6.77 (1H, m), 6.94 (2H, d, J = 2.3 Hz), 8.26 (1H, s). m/z[M + H]⁺490.2 19

1H-NMR (DMSO-d6) δ: 0.94 (6H, d, J = 6.6 Hz), 2.09 (3H, s), 2.73-2.80(1H, m), 3.13 (2H, d, J = 5.1 Hz), 3.78 (6H, s), 4.24-4.29 (1H, m),4.40-4.45 (1H, m), 4.53-4.57 (1H, m), 4.70-4.74 (1H, t, J = 8.4 Hz),5.69-5.73 (1H, m), 6.17 (1H, d, J = 15.4 Hz), 6.60-6.67 (2H, m), 6.95(2H, d, J = 2.2 Hz), 8.32 (1H, s). m/z[M + H]⁺ 490.2 20

1H-NMR (DMSO-d6) δ: 1.36-1.50 (6H, m), 2.32 (4H, brs), 3.05 (2H, d, J =6.2 Hz), 3.78 (6H, s), 4.24-4.30 (1H, m), 4.42 (1H, t, J = 9.5 Hz),4.53-4.57 (1H, m), 4.72 (1H, t, J = 8.8 Hz), 5.68-5.74(1H, m), 6.16 (1H,d, J = 15.4 Hz), 6.60-6.68 (2H, m) 6.94 (2H, d, J = 2.2 Hz), 8.29 (1H,s). m/z[M + H]⁺ 502.2

TABLE 6 21

1H-NMR (DMSO-d6) δ: 2.31-2.39 (4H, m), 3.10 (2H, d, J = 5.5 Hz),3.50-3.60 (4H, m), 3.78 (6H, s), 4.23-4.31 (1H, m), 4.43 (1H, t, J = 9.2Hz), 4.45-4.58 (1H, m), 4.73 (1H, t, J = 8.8 Hz), 5.67-5.75 (1H, m),6.20 (1H, d, J = 15.0 Hz), 6.61-6.70 (2H, m), 6.94 (2H, d, J = 1.8 Hz),8.26 (1H, s). m/z[M + H]⁺ 504.1 22

1H-NMR (DMSO-d6) δ: 1.75-1.95 (1H, m), 2.07- 2.15 (1H, m), 2.32-2.83(4H, m), 3.20-3.25 (2H, m), 3.78 (6H, s), 4.25-4.30 (1H, m), 4.43 (1H,t, J = 9.3 Hz), 4.55-4.60 (1H, m), 4.73 (1H, t, J = 8.8 Hz), 5.09-5.13(0.5H, m), 5.22-5.27 (0.5H, m), 5.69-5.73 (1H, m), 6.19 (1H, d, J = 15.4Hz), 6.60-6.72 (2H, m), 6.95 (2H, d, J = 2.2 Hz), 8.37 (1H, s). m/z[M +H]⁺ 506.1 23

1H-NMR (DMSO-d6) δ: 1.75-1.95 (1H, m), 2.07- 2.15 (1H, m), 2.32-2.83(4H, m), 3.20-3.25 (2H, m), 3.78 (6H, s), 4.25-4.30 (1H, m), 4.43 (1H,t, J = 9.3 Hz), 4.55-4.60 (1H, m), 4.73 (1H, t, J = 8.8 Hz), 5.09-5.13(0.5H, m), 5.22-5.27 (0.5H, m), 5.69-5.73 (1H, m), 6.19 (1H, d, J = 15.4Hz), 6.60-6.72 (2H, m), 6.95 (2H, d, J = 2.2 Hz), 8.37 (1H, s). m/z[M +H]⁺ 506.1

TABLE 7 24

1H-NMR (DMSO-d6) δ: 1.31 (6H, t, J = 7.0 Hz), 4.04 (4H, q, J = 7.0 Hz),4.30 (1H, dd, J = 10.5, 5.4 Hz), 4.41-4.48 (1H, m), 4.58 (1H, dd, J =9.3, 5.4 Hz), 4.71-4.78 (1H, m), 5.68-5.76 (2H, m), 6.16 (1H, dd, J =17.0, 2.1 Hz), 6.38 (1H, dd, J = 17.0, 10.4 Hz), 6.57 (1H, t, J = 2.2Hz), 6.90 (2H, d, J = 2.2 Hz), 8.26 (1H, s). m/z[M + H]⁺ 433.2 25

1H-NMR (DMSO-d6) δ: 1.31 (6H, t, J = 7.1 Hz), 2.14 (6H, s), 3.00-3.05(2H, m), 4.04 (4H, q, J = 7.1 Hz), 4.27 (1H, dd, J = 10.4, 5.0 Hz),4.39-4.46 (1H, m), 4.55 (1H, dd, J = 9.3, 5.4 Hz), 4.68-4.76 (1H, m),5.66-5.75 (1H, m), 6.13-6.20 (1H, m), 6.54-6.69 (2H, m), 6.87-6.91 (2H,m), 8.25 (1H, s). m/z[M + H]⁺ 490.2 26

1H-NMR (DMSO-d6) δ: 1.57-1.92 (2H, m), 2.64- 2.85 (1H, m), 3.10-3.65(4H, m), 3.72 (6H, s), 4.28 4.35 (2H, m), 5.53-5.59 (1H, m), 5.95-6.06(1H, m), 6.38-6.55 (2H, m), 6.83-6.86 (2H, m), 8.16-8.19 (1H, m).m/z[M + H]⁺ 433.1 27

1H-NMR (DMSO-d6) δ: 1.55-2.32 (4H, m), 2.87- 4.71 (5H, m), 3.78(6H, m),5.50 5.75 (1H, m), 6.05-6.20 (1H, m), 6.61 (1H, t, J = 2.2 Hz),6.65-6.91 (1H, m), 6.91 (2H, d, J = 2.2 Hz), 8.27 (1H, s). m/z[M + H]⁺433.1

TABLE 8 28

1H-NMR (DMSO-d6) δ: 2.48-2.50 (2H, m), 3.27- 4.35 (11H, m), 4.89 (0.5H,t, J = 5.7 Hz), 5.02 (0.5H, t, J = 5.7 Hz), 5.28 5.37 (1H, m), 5.66-5.71 (1H, m), 6.13-6.20 (1H, m), 6.60 (1H, t, J = 2.3 Hz), 6.93 (2H, d,J = 2.3 Hz), 8.26 (1H, s). m/z[M + H]⁺ 449.1 29

1H-NMR (CDCl3) δ: 3.20-3.35 (1H, m), 3.81 (6H, s), 3.91-4.02 (1H, m),4.14-4.38 (3H, m), 4.58 4.70 (2H, m), 5.66 (1H, d, J = 10.5 Hz), 5.85-6.05 (2H, m), 6.16 (1H, dd, J = 17.0, 10.4 Hz), 6.33 (1H, d, J = 17.1Hz), 6.54 (1H, s), 6.74 (2H, dd, J = 3.3, 2.6 Hz), 8.35 (1H, s). m/z[M +H]⁺ 419.1 30

m/z[M + H]⁺ 419.2 31

1H-NMR (CDCl3) δ: 2.08-2.10(2H, m), 2.20- 2.30(4H, m), 2.81-2.98(1H, m),3.22-3.40(1H, m), 3.82(6H, s), 4.15-4.25(1H, m), 4.80-4.88(1H, m)4.96-5.04(1H, m), 5.72(1H, dd, J = 10.8, 2.0 Hz), 6.30(1H, dd, J = 16.8,2.0 Hz), 6.54(1H, t, J = 2.0 Hz) 6.61(1H, dd, J = 17.2, 10.8 Hz),6.73(2H, d, J = 2.4 Hz), 8.42(1H, s) m/z[M + H]⁺ 433.2

TABLE 9 32

1H-NMR (CDCl3) δ: 2.34-2.42 (1H, m), 2.80-3.10 (2H, m), 3.82 (6H, s),4.05-4.20 (2H, m), 4.20- 4.50 (1H, m), 4.70-4.95 (1H, m), 5.30-5.40 (1H,m), 5.75-5.80 (1H, m), 5.98 (2H, brs), 6.40- 6.50 (1H, m), 6.54 (1H, s),6.70-6.73 (2H, m), 8.38 (1H, m). m/z [M + H]⁺ 443.1 33

m/z [M + H]⁺ 447.2 34

1H-NMR (DMSO-d6) δ: 3.77 (6H, s), 5.50 (2H, s), 5.72 (1H, dd, J = 10.2,2.1 Hz), 6.22 (1H, dd, J = 17.1, 2.1 Hz), 6.38 (1H, dd, J = 17.1, 10.2Hz), 6.59 (1H, t, J = 2.3 Hz), 6.90 (2H, d, J = 2.3 Hz), 7.00 (1H, d, J= 7.8 Hz), 7.29 (1H, t, J = 7.8 Hz), 7.45 (1H, s), 7.63-7.66 (1H, m),8.28 (1H, s), 10.12 (1H, s). m/z [M + H]⁺ 455.2 35

m/z [M + H]⁺ 476.2 36

1H-NMR (CDCl3) δ: 2.09 (2H, d, J = 10.7 Hz), 2.29 (8H, m), 2.80-2.98(1H, m), 3.12 (2H, d, J = 4.9 Hz), 3.19-3.40 (1H, m), 3.81 (6H, s),4.12-4.32 (1H, m), 4.74-4.90 (1H, m), 4.94-4.96 (1H, m), 5.74-5.85 (2H,m), 6.43-6.55 (2H, m), 6.73 (2H, s), 6.80-6.80 (1H, m), 8.37 (1H, s).m/z [M + H]⁺ 490.2

TABLE 10 37

m/z [M + H]⁺ 504.2 38

1H-NMR (DMSO-d6) δ: 2.13 (3H, s), 2.17 (3H, s), 2.29-2.49 (2H, m), 3.01(1H, d, J = 6.3 Hz), 3.06 (1H, d, J = 6.3 Hz), 3.40-4.10 (4H, m), 3.76(6H, m), 5.23-5.37 (1H, m), 6.36 (0.5H, d, J = 15.0 Hz), 6.43 (0.5H, d,J = 15.0 Hz), 6.54 (1H, t, J = 2.2 Hz), 6.60-6.69 (1H, m), 6.74 (2H, d,J = 2.2 Hz), 7.71 (0.5H, s), 7.76 (0.5H, s), 8.16 (1H, t, J = 2.2 Hz).m/z [M + H]⁺ 475.1 39

1H-NMR (DMSO-d6) δ: 2.30-2.50 (2H, m), 3.40- 4.15 (4H, m), 3.77 (6H, m),5.20-5.40 (1H, m), 5.60-5.80 (1H, m), 6.10-6.20 (1H, m), 6.54 (1H, s),6.54-6.74 (1H, m), 6.74 (2H, s), 7.72 (0.5H, s), 7.76 (0.5H, s), 8.18(1H, s), m/z [M + H]⁺ 418.0 40

1H-NMR (DMSO-d6) δ: 1.62-1.72 (4H, m), 2.28- 2.51 (6H, m), 3.17 (1H, d,J = 5.9 Hz), 3.21 (1H, d, J = 5.9 Hz), 3.52-3.90 (9.5H, m), 4.02- 4.12(0.5H, m), 5.20-5.35 (1H, m), 6.30-6.45 (1H, m), 6.53 (1H, t, J = 2.4Hz), 6.60-6.80 (3H, m), 7.71 (0.5H, s), 7.75 (0.5H, s), 8.17 (0.5H, s),8.18 (0.5H, m). m/z [M + H]⁺ 501.1

TABLE 11 41

1H-NMR (DMSO-d6) δ : 2.15 (1.5H, s), 2.17 (1.5H, s), 2.18-2.42 (10H, m),3.05 (1H, d, J = 6.2 Hz), 3.10 (1H, d, J = 6.2 Hz), 3.47-3.94 (9.5H, m),4.02-4.10 (0.5H, m), 5.22-5.38 (1H, m), 6.32-6.45 (1H, m), 6.54 (1H, d,J = 1.8 Hz), 6.57-6.70 (1H, m), 6.74 (2H, d, J = 1.5 Hz), 7.70 (0.5H,s), 7.75 (0.5H, s), 8.14-8.17 (1H, m). m/z [M + H]⁺ 530.2 42

1H-NMR (DMSO-d6) δ : 1.32-1.42 (2H, m), 1.60- 1.75 (2H, m), 1.95-2.06(2H, m), 2.29-2.52 (2H, m), 2.60-2.70 (2H, m), 3.04 (1H, d, J = 6.2 Hz),3.08 (1H, d, J = 6.2 Hz), 3.35-3.94 (10.5H, m), 4.05-4.10 (0.5H, m),5.23-5.37 (1H, m), 6.34 (0.5H, d, J = 15.4 Hz), 6.41 (0.5H, d, J = 15.4Hz), 6.53 (1H, t, J = 2.4 Hz), 6.59-6.68 (1H, m), 6.74 (2H, d, J = 1.5Hz), 7.71 (0.5H, s), 7.75 (0.5H, s), 8.15-8.17 (1H, m). m/z [M + H]⁺531.1 43

1H-NMR (DMSO-d6) δ : 1.60-1.90 (4H, m), 2.20- 2.50 (6H, m), 3.07 (1H, d,J = 6.1 Hz), 3.12 (1H, d, J = 6.1 Hz), 3.52-3.89 (9.5H, m), 4.03- 4.10(0.5H, m), 4.55 4.80 (1H, m), 5.20-5.40 (1H, m), 6.35-6.45 (1H, m), 6.53(1H, t, J = 2.2 Hz), 6.55-6.70 (1H, m), 6.74 (2H, d, J = 2.2 Hz), 7.71(0.5H, s), 7.76 (0.5H, s), 8.17 (0.5H, s), 8.18 (0.5H, m). m/z [M + H]⁺533.1

TABLE 12 44

1H-NMR (DMSO-d6) δ : 2.18-2.54 (2H, m), 2.66- 2.93 (4H, m), 3.18-3.24(2H, m), 3.65-3.93 (9.5H, m), 4.04-4.10 (0.5H, m), 5.24-5.38 (1H, m),6.35-6.50 (1H, m), 6.53 (3H, t, J = 2.2 Hz), 6.59-6.69 (1H, m), 6.74(2H, d, J = 2.2 Hz), 7.71 (0.5H, s), 7.76 (0.5H, s), 8.17 (0.5H, s),8.18 (0.5H, m). m/z [M + H]⁺ 537.1 45

1H-NMR (DMSO-d6) δ : 1.88-2.02 (4H, m), 2.42- 2.54 (6H, m), 3.15 (1H, d,J = 6.1 Hz), 3.20 (1H, d, J = 6.1 Hz), 3.49-3.92 (9.5H, m), 4.05- 4.10(0.5H, m), 5.22-5.38 (1H, m), 6.38-6.48 (1H, m), 6.54 (1H, t, J = 2.4Hz), 6.60-6.70 (1H, m), 6.74 (2H, d, J = 2.4 Hz), 7.71 (0.5H, s), 7.76(0.5H, s), 8.17 (0.5H, s), 8.18 (0.5H, m). m/z [M + H]⁺ 551.1 46

1H-NMR (DMSO-d6) δ : 1.97 (1.5H, s), 2.04 (1.5H, s), 2.37-2.43 (2H, m),2.97-3.89 (9.5H, m), 4.05-4.12 (0.5H, m), 5.25-5.35 (1H, m), 6.54 (1H,d, J = 2.4 Hz), 6.74 (2H, d, J = 2.4 Hz), 7.74 (0.5H, s), 7.76 (0.5H,s), 8.16 (0.5H, s), 8.17 (0.5H, s). m/z [M + H]⁺ 430.1 47

1H-NMR (DMSO-d6) δ : 1.30 (3H, s), 1.37 (3H, s), 2.30-2.50 (2H, s),3.06-4.10 (4H, m), 3.70 (6H, s), 5.20-5.35 (1H, m), 5.56 (0.5H, s), 5.64(0.5H, s), 6.47 (1H, s), 6.68 (2H, d, J = 2.2 Hz), 7.70 (0.5H, s), 7.73(0.5H, s), 8. 8.10 (0.5H, s), 8.11 (0.5H, m). m/z [M + H]⁺ 474.4

TABLE 13 48

1H-NMR (DMSO-d6) δ : 1.76-2.85 (8H, m), 3.16- 4.08 (12H, m), 5.05-5.38(2H, m), 6.35-6.48 (1H, m), 6.53 (1H, t, J = 2.3 Hz), 6.62-6.72 (1H, m),6.74 (2H, d, J = 2.3 Hz), 7.71 (0.5H, s), 7.76 (0.5H, s), 8.17 (0.5H,s), 8.18 (0.5H, m). m/z [M + H]⁺ 519.1 49

1H-NMR (DMSO-d6) δ : 1.76-2.85 (8H, m), 3.16- 4.08 (12H, m), 5.05-5.38(2H, m), 6.35-6.48 (1H, m), 6.53 (1H, t, J = 2.3 Hz), 6.62-6.72 (1H, m),6.74 (2H, d, J = 2.3 Hz), 7.71 (0.5H, s), 7.76 (0.5H, s), 8.17 (0.5H,s), 8.18 (0.5H, m). m/z [M + H]⁺ 519.1 50

1H-NMR (DMSO-d6) δ : 1.20 1.50 (6H, m), 2.19- 2.43 (6H, m), 2.96 (1H, d,J = 6.2 Hz), 3.00 (1H, d, J = 5.5 Hz), 3.31-4.02 (4H, m), 3.73 (6H, s),5.15-5.30 (1H, m), 6.31 (1H, dd, J = 28.7, 15.0 Hz), 6.47 (1H, d, J =2.1 Hz), 6.50- 6.65 (1H, m), 6.66 (2H, d, J = 2.1 Hz), 7.63 (0.5H, s),7.68 (0.5H, s), 8.10 (1H, s). m/z [M + H]⁺ 515.1 51

1H-NMR (DMSO-d6) δ : 3.76 (6H, s), 4.20-4.27 (1H, m), 4.30-4.39 (1H, m),4.50-4.60 (1H, m), 4.62-4.68 (1H, m), 5.50-5.60 (1H, m), 5.64 (1H, dd, J= 10.3, 2.3 Hz), 6.08 (1H, dd, J = 16.9, 2.3 Hz), 6.29 (1H, dd, J =16.9, 10.3 Hz), 6.47 (1H, t, J = 2.1 Hz), 6.67 (2H, d, J = 2.1 Hz), 7.98(1H, s), 8.09 (1, s). m/z [M + H]⁺ 404.0

TABLE 14 52

1H-NMR (DMSO-d6) δ : 2.72 (6H, s), 3.76 (6H, s), 3.84 (2H, d, J = 6.8Hz), 4.30-4.35 (1H, m), 4.44 (1H, t, J = 9.5 Hz), 4.60-4.75 (2H, m),5.50-5.60 (1H, m), 6.42 (1H, d, J = 15.4 Hz), 6.54 (1H, t, J = 2.4 Hz),6.59-6.67 (1H, m), 6.73 (2H, d, J = 2.4 Hz), 8.03 (1H, s), 8.15 (1H, s).m/z [M + H]⁺ 461.1 53

1H-NMR (DMSO-d6) δ : 1.69 (4H, brs), 2.35- 2.55 (4H, m), 3.22 (2H, brs),3.76 (6H, s), 4.25-4.45 (2H, m), 4.55-4.75 (2H, m), 5.49-5.59 (1H, t,m), 6.15 (1H, d, J = 15.5 Hz), 6.54 (1H, t, J = 2.3 Hz), 6.66 (1H, dt, J= 15.5, 6.0 Hz), 6.74 (2H, d, J = 2.3 Hz), 8.05 (1H, s), 8.15 (1H, s).m/z [M + H]⁺ 487.1 54

1H-NMR (DMSO-d6) δ : 2.01 (3H, s), 3.77 (6H, s), 4.25-4.41 (1H, m), 4.38(1H, t, J = 9.3 Hz), 4.52-4.63 (2H, m), 5.52-5.57 (1H, m), 6.54 (1H, t,J = 2.2 Hz), 6.74 (2H, d, J = 2.2 Hz), 8.03 (1H, s), 8.16 (1H, s). m/z[M + H]⁺ 416.4 55

1H-NMR (DMSO-d6) δ : 1.94-2.00 (2H, m), 3.11- 3.15 (6H, t, J = 7.0 Hz),3.77 (6H, s), 4.25 4.28 (1H, m), 4.39 (1H, t, J = 9.3 Hz), 4.58- 4.75(2H, m), 5.50-5.58 (1H, m), 6.07 (1H, d, J = 15.4 Hz), 6.50-6.58 (2H,m), 6.74 (2H, d, J = 2.6 Hz), 8.05 (1H, s), 8.15 (1H, s). m/z [M + H]⁺473.1

TABLE 15 56

1H-NMR (DMSO-d6) δ : 0.98 (3H, t, J = 7.1 Hz), 2.13 (3H, s), 2.35 (2H,q, J = 7.2 Hz), 3.10 (2H, d, J = 6.2 Hz), 3.77 (6H, s), 4.24-4.29 (1H,m), 4.40 (1H, t, J = 9.5 Hz), 4.65-4.75 (2H, m), 5.53-5.57 (1H, m), 6.15(1H, d, J = 15.4 Hz), 6.54 (1H, t, J = 2.5 Hz), 6.64 (1H, dt, J = 15.4,6.1 Hz), 6.74 (2H, d, J = 2.5 Hz), 8.05 (1H, s), 8.15 (1H, s). m/z [M +H]⁺ 475.1 57

1H-NMR (DMSO-d6) δ : 0.95 (3H, d, J = 6.6 Hz), 1.14 (3H, d, J = 6.6 Hz),3.05-3.15 (1H, m), 3.64 (2H, d, J = 5.9 Hz), 3.77 (6H, s), 4.22- 4.70(4H, m), 5.50 5.60 (1H, m), 6.25-6.36 (1H, m), 6.52-6.57 (1H, m),6.63-6.70 (1H, m), 6.73- 6.79 (2H, m), 8.02 (0.5H, s), 8.04 (0.5H, s),8.16 (1H, s). m/z [M + H]⁺ 475.1 58

1H-NMR (DMSO-d6) δ : 0.94 (6H, d, J = 6.6 Hz), 2.09 (3H, s), 2.74-2.80(1H, m), 3.13 (2H, d, J = 5.1 Hz), 3.77 (6H, s), 4.24-4.29 (1H, m), 4.40(1H, t, J = 9.2 Hz), 4.63-4.72 (1H, m), 5.53-5.57 (1H, m), 6.15 (1H, d,J = 15.3 Hz), 6.54 (1H, t, J = 2.2 Hz), 6.62 (2H, dt, J = 15.3, 6.0 Hz),6.74 (2H, d, J = 2.2 Hz), 8.05 (1H, s), 8.15 (1H, s). m/z [M + H]⁺ 489.2

TABLE 16 59

1H-NMR (DMSO-d6) δ : 0.95 (6H, t, J = 7.1 Hz), 2.45 (4H, q, J = 7.1 Hz),3.18 (2H, d, J = 5.9 Hz), 3.76 (6H, s), 4.25-4.29 (1H, m), 4.40 (1H, t,J = 9.5 Hz), 4.57-4.59 (1H, m), 4.68 (1H, t, J = 8.8 Hz), 5.52-5.70 (1H,m), 6.17 (1H, d, J = 15.0 Hz), 6.53-6.56 (1H, m), 6.66 (1H, dt, J =15.0, 6.1 Hz), 6.74-6.75 (2H, m), 8.06 (1H, s), 8.15 (1H, s). m/z [M +H]⁺ 489.2 60

1H-NMR (DMSO-d6) δ : 2.18 (3H, s), 3.15 (2H, d, J = 5.5 Hz), 3.22 (3H,s), 3.77 (6H, s), 4.25- 4.30 (1H, m), 4.40 (1H, t, J = 9.7 Hz), 4.55-4.75 (2H, m), 5.51-5.59 (1H, m), 6.17 (1H, d, J = 15.4 Hz), 6.54 (1H, t,J = 2.4 Hz), 6.63 (1H, dt, J = 15.4, 6.0 Hz), 6.74 (2H, d, J = 2.4 Hz),8.05 (1H, s), 8.15 (1H, s). m/z [M + H]⁺ 505.1 61

1H-NMR (DMSO-d6) δ : 1.30-1.42 (2H, m), 1.65- 1.75 (2H, m), 2.03 (2H, t,J = 10.3 Hz), 2.62- 2.70 (2H, m), 3.06 (2H, d, J = 5.1 Hz), 3.77 (6H,s), 4.22-4.30 (1H, m), 4.40 (1H, t, J = 9.2 Hz), 4.55-4.72 (2H, m),5.52-5.58 (1H, m), 6.13 (1H, d, J = 15.0 Hz), 6.54 (1H, t, J = 2.3 Hz),6.58-6.65 (1H, m), 6.74 (2H, d, J = 2.3 Hz), 8.06 (1H, s), 8.15 (1H, s).m/z [M + H]⁺ 517.1

TABLE 17 62

1H-NMR (DMSO-d6) δ : 1.48-1.58 (1H, m), 1.92- 2.00 (1H, m), 2.31-2.71(4H, m), 3.19 (2H, brs), 3.77 (6H, s), 4.15-4.30 (2H, m), 4.35- 4.44(1H, m), 4.55-4.72 (2H, m), 5.50-5.60 (1H, m), 6.15 (1H, d, J = 15.4Hz), 6.54 (1H, t, J = 2.2 Hz), 6.60-6.70 (1H, m), 6.75 (2H, d, J = 2.2Hz), 8.05 (1H, s), 8.15 (1H, d, J = 1.8 Hz). m/z [M + H]⁺ 503.1 63

1H-NMR (DMSO-d6) δ : 1.48-1.58 (1H, m), 1.92- 2.00 (1H, m), 2.31-2.71(4H, m), 3.19 (2H, brs), 3.77 (6H, s), 4.15-4.30 (2H, m), 4.35- 4.44(1H, m), 4.55-4.72 (2H, m), 5.50-5.60 (1H, m), 6.15 (1H, d, J = 15.4Hz), 6.54 (1H, t, J = 2.2 Hz), 6.60-6.70 (1H, m), 6.75 (2H, d, J = 2.2Hz), 8.05 (1H, s), 8.15 (1H, d, J = 1.8 Hz). m/z [M + H]⁺ 503.1 64

1H-NMR (CDCl3) δ : 1.67-2.22 (4H, m), 2.71-2.95 (1H, m), 3.19-3.35 (1H,m), 3.81 (6H, s), 4.09- 4.25 (1H, m), 4.83-4.95 (2H, m), 5.61-5.78 (3H,m), 6.28 6.68 (5H, m), 7.24 (1H, s), 8.31 (1H, s). m/z [M + H]⁺ 432.2 65

1H-NMR (CDCl3) δ : 1.65-2.00 (4H, m), 2.29 (6H, s), 2.77-2.92 (1H, m),3.12 (2H, d, J = 5.9 Hz), 3.22-3.38 (1H, m), 3.81 (6H, s), 4.14-4.30(1H, m), 4.85-4.97 (2H, m), 5.66 (2H, br s), 6.44-6.55 (2H, m), 6.65(2H, d, J = 2.2 Hz), 6.84-6.95 (1H, m), 7.24 (1H, s), 8.31 (1H, s). m/z[M + H]⁺ 489.2

TABLE 18 66

1H-NMR (CDCl3) δ : 2.37-2.48 (1H, m), 2.82- 2.93 (1H, m), 3.41-3.52 (1H,m), 3.82 (9H, s), 3.86-3.95 (1H, m), 4.25-4.38 (1H, m), 4.69 (1H, t, J =8.3 Hz), 5.43-5.56 (1H, m), 5.65-5.83 (2H, m), 6.36-6.50 (3H, m), 6.65(2H, d, J = 2.0 Hz), 7.37 (1H, s), 8.30 (1H, s). m/z [M + H]⁺ 476.2 67

m/z [M + H]⁺ 475.2 68

1H-NMR (CDCl3) δ : 2.75-2.95 (1H, m), 2.99-3.15 (1H, m), 3.82 (6H, s),4.00 (1H, t, J = 9.8 Hz), 4.32-4.50 (1H, m), 5.44-5.85 (5H, m), 6.44(2H, d, J = 5.9 Hz), 6.49 (1H, t, J = 2.2 Hz), 6.66 (2H, d, J = 2.2 Hz),7.44 (1H, br s), 8.30 (1H, s), 8.10 (1H, s). m/z [M + H]⁺ 486.1 69

m/z [M + H]⁺ 500.2 70

1H-NMR (CDCl3) δ : 2.36 (6H, br s), 2.69-2.90 (1H, m), 2.94-3.10 (1H,m), 3.69-3.80 (2H, m), 3.82 (6H, s), 3.92-4.05 (1H, m), 4.32-4.47 (1H,m), 5.39-5.83 (5H, m), 6.35-6.47 (2H, m), 6.49 (1H, t, J = 2.1 Hz), 6.65(2H, d, J = 2.2 Hz), 7.44 (1H, br s), 8.30 (1H, s). m/z [M + H]⁺ 543.2

TABLE 19 71

1H-NMR (CDCl3) δ : 2.30 (6H, d, J = 7.6 Hz), 2.35-2.68 (3H, m),2.77-2.92 (1H, m), 3.14 (3H, d, J = 79.5 Hz), 3.31-3.47 (1H, m),3.69-3.81 (1H, m), 3.82 (6H, s), 3.95 (2H, q, J = 9.9 Hz), 4.35 (1H, t,J = 8.9 Hz), 5.08 (1H, t, J = 7.3 Hz), 5.50-5.79 (4H, m), 6.37-6.52 (3H,m), 6.65 (2H, t, J = 2.2 Hz), 7.59 (1H, d, J = 28.0 Hz), 8.30 (1H, s).m/z [M + H]⁺ 546.3 72

m/z [M + H]⁺ 503.3 73

1H-NMR (DMSO-d6) δ : 2.21-2.44 (2H, m), 3.34- 3.87 (9.5H, m), 4.00-4.10(0.5H, m), 5.02-5.18 (1H, m), 5.58-5.68 (1H, m), 5.88 (2H, brs),6.05-6.18 (1H, m), 6.45-6.65 (4H, m), 6.82-6.88 (1H, m), 7.60-7.63 (2H,m). m/z [M + H]⁺ 417.5

TABLE 20 Ref. Ex . Comp. Structural formula Physical properties 1

m/z [M + H]⁺ 441.2 2

m/z [M + H]⁺ 289.1 3

m/z [M + H]⁺ 421.2 4

m/z [M + H]⁺ 500.3 5

m/z [M + H]⁺ 446.2 6

m/z [M + H]⁺ 473.2 7

m/z [M + H]⁺ 417.2

Test Example 1

Measurement of Inhibitory Effect on FGFR2 Kinase Activity

When setting conditions for the measurement of the inhibitory effect ofthe compounds on FGFR2 kinase activity, FL-Peptide 22 (Caliper LifeSciences, Inc.) was used as a substrate. The purified recombinant humanFGFR2 protein used in the test was purchased from Carna Biosciences,Inc. In the measurement of the inhibitory effect of the compounds,first, a test compound was gradually diluted with dimethylsulfoxide(DMSO) to a concentration that was 20 times higher than the finalconcentration. Next, the purified human FGFR2 protein, FL-Peptide 22(final concentration: 1.5 μM), magnesium chloride (final concentration:5 mM), ATP (final concentration: 75 μM), and the test compound DMSOsolution (final concentration of DMSO: 5%) were added to a reactionbuffer (15 mM Tris-HCl pH 7.5, 0.01% Tween-20, 2 mM DTT), and themixture was incubated at 25° C. for 120 minutes to perform a kinasereaction. EDTA (final concentration: 30 mM) diluted with a separationbuffer (Caliper Life Sciences, Inc.) was added thereto to terminate thekinase reaction. Finally, using a LabChip (registered trademark) 3000system (Caliper Life Sciences, Inc.; excitation wavelength: 488 nm,detection wavelength: 530 nm), phosphorylated peptides andnon-phosphorylated peptides were separated, and the amount of eachpeptide was measured. The level of phosphorylation was determined fromthe quantitative ratio. The compound concentration at whichphosphorylation was inhibited by 50% was defined as the IC₅₀ value (nM).Table 21 shows the results.

The results demonstrated that all of the compounds of the presentinvention, represented by the test compounds, which had a dialkoxybenzene ethynyl group and a partial structure of α,β-unsaturated amide,exhibited a high FGFR2 inhibitory effect. Conversely, the compounds ofthe Reference Examples, which did not have a dialkoxy benzene ethynylgroup or a partial structure of α,β unsaturated amide, showed aremarkably lower FGFR2 inhibitory effect.

TABLE 21 IC₅₀ value Test compound (nM) Ex. Compound 1 2.4 Ex. Compound 21.1 Ex. Compound 5 1.1 Ex. Compound 8 6.9 Ex. Compound 9 1.1 Ex.Compound 10 1.2 Ex. Compound 12 2.2 Ex. Compound 14 3.8 Ex. Compound 153.0 Ex. Compound 16 3.8 Ex. Compound 17 4.1 Ex. Compound 18 4.3 Ex.Compound 19 2.7 Ex. Compound 20 3.3 Ex. Compound 22 5.6 Ex. Compound 236.8 Ex. Compound 24 7.2 Ex. Compound 28 3.7 Ex. Compound 29 9.3 Ex.Compound 32 0.4 Ex. Compound 38 4.7 Ex. Compound 39 0.4 Ex. Compound 403.2 Ex. Compound 42 6.6 Ex. Compound 46 4.3 Ex. Compound 47 2.0 Ex.Compound 48 6.8 Ex. Compound 49 7.0 Ex. Compound 50 5.8 Ex. Compound 51<0.3 Ex. Compound 52 1.0 Ex. Compound 53 0.6 Ex. Compound 55 2.5 Ex.Compound 56 1.5 Ex. Compound 57 1.5 Ex. Compound 59 1.2 Ex. Compound 601.5 Ex. Compound 61 2.3 Ex. Compound 63 1.0 Ex. Compound 64 7.9 Ex.Compound 65 7.0 Ex. Compound 66 <0.3 Ex. Compound 68 1.2 Ex. Compound 698.8 Ex. Compound 73 4.9 Ref. Ex. Compound 1 280 Ref. Ex. Compound 2 270Ref. Ex. Compound 3 190 Ref. Ex. Compound 4 >10000 Ref. Ex. Compound 5190 Ref. Ex. Compound 6 110 Ref. Ex. Compound 7 1600

Test Example 2 Cell Growth-Inhibitory Effect on Human-Derived GastricCancer Cell Line with High Expression of FGFR

Human-derived gastric cancer OCUM-2MD3 cells, which overexpressed FGFR2,were subcultured daily in a Dulbecco's modified Eagle's medium (DMEM)containing 10% fetal bovine serum (FBS) at a cell density of not morethan 80%. In order to initiate a test of the cell growth-inhibitoryeffect of the compounds, the OCUM-2MD3 cells were suspended in the aboveDMEM medium and seeded in a 96-well flat-bottom plate so that each wellcontained 3,000 cells. Then, the cells were cultured in an incubatorcontaining 5% carbon dioxide gas at 37° C. for one day. On the followingday, the test compound was gradually diluted with DMSO to aconcentration 100 times higher than the final concentration. The DMSOsolution of the test compound was diluted with the medium used forcultivation, and the diluted solution was added to each well of the cellculture plate so that the final concentration of DMSO became 0.5%. Then,the cells were cultured in an incubator containing 5% carbon dioxide gasat 37° C. for 72 hours. The number of cells was measured at the time ofthe addition of the test compound and 72 hours later after culture byusing a Cell Counting Kit-8 (produced by Dojindo Laboratories) accordingto a protocol recommended by Dojindo Laboratories. The reagent of thekit was added to each plate, and a color reaction was performed in anincubator containing 5% carbon dioxide gas at 37° C. for a predeterminedtime. After completion of the reaction, the absorbance at a wavelengthof 450 nm was measured by a microplate reader. The cell-growthinhibition rate was calculated by the following formula, and theconcentration of the test compound at which the cell growth wasinhibited by 50% (GI₅₀ (nM)) was determined. Table 22 shows the results.

The results demonstrated that all of the compounds of the presentinvention, represented by the test compounds, showed a high growthinhibitory effect on the human-derived gastric cancer OCUM-2MD3 cellline.Growth inhibition rate (%)=(C−T)/(C−C0)×100

T: absorbance of well to which test compound was added

C: absorbance of well to which test compound was not added

C0: absorbance of well measured before addition of compound

TABLE 22 Test compound GI₅₀ (nM) Ex. Compound 2 <4.6 Ex. Compound 4 17Ex. Compound 8 5 Ex. Compound 12 <4.6 Ex. Compound 15 <4.6 Ex. Compound17 4.6 Ex. Compound 19 <4.6 Ex. Compound 24 13 Ex. Compound 25 13 Ex.Compound 28 5 Ex. Compound 29 20 Ex. Compound 32 <4.6 Ex. Compound 39<4.6 Ex. Compound 40 7 Ex. Compound 64 12 Ex. Compound 65 13 Ex.Compound 66 <4.6 Ex. Compound 68 8 Ex. Compound 73 7

Test Example 3

Measurement of Inhibitory Effect on FGFR1 Kinase Activity

When setting conditions for the measurement of the inhibitory effect ofthe compounds on FGFR1 kinase activity, a biotinylated peptide(biotin-EEPLYWSFPAKKK) was synthesized for use as a substrate byutilizing the amino acid sequence of FL-Peptide 22 (Caliper LifeSciences, Inc.) with biotin. The purified recombinant human FGFR1protein used in the test was purchased from Carna Biosciences, Inc. Inthe measurement of the inhibitory effect of the compounds, first, a testcompound was gradually diluted with dimethylsulfoxide (DMSO) to aconcentration 20 times higher than the final concentration. Next, thepurified human FGFR1 protein, substrate peptide (final concentration:250 nM), magnesium chloride (final concentration: 5 mM), ATP (finalconcentration: 190 μM), and the test compound DMSO solution (finalconcentration of DMSO: 5%) were added to a reaction buffer (15 mMTris-HCl pH 7.5, 0.01% Tween-20, 2 mM DTT), and the mixture wasincubated at 25° C. for 120 minutes to perform a kinase reaction. EDTAwas added thereto to a final concentration of 40 mM to thereby terminatethe reaction. Then, a detection solution containing Eu-labeledanti-phosphorylated tyrosine antibody PT66 (PerkinElmer) and SureLightAPC-SA (PerkinElmer) was added, and the resulting mixture was allowed tostand at room temperature for 2 hours or more. Finally, the intensity offluorescence when excitation light with a wavelength of 337 nm wasirradiated was measured by a PHERAstar FS (BMG LABTECH) at twowavelengths of 620 nm and 665 nm. The amount of phosphorylation wasdetermined from the fluorescence intensity ratio of the two wavelengths.The compound concentration at which phosphorylation was inhibited by 50%was defined as the IC₅₀ value (nM). Table 23 below shows the results.

Test Example 4

Measurement of Inhibitory Effect on FGFR3 Kinase Activity

The inhibitory effect of the compounds on FGFR3 kinase activity wasmeasured according to the method of Test Example 3. Purified recombinanthuman FGFR3 protein was purchased from Carna Biosciences, Inc. The finalconcentration of ATP was 50 μM. Table 23 shows the results.

Test Example 5

Measurement of Inhibitory Effect on FGFR4 Kinase Activity

The inhibitory effect of the compounds on FGFR4 kinase activity wasmeasured according to the method of Test Example 3. Purified recombinanthuman FGFR4 protein was purchased from Carna Biosciences, Inc. The finalconcentration of ATP was 200 μM. Table 23 shows the results.

The results of Test Examples 3 to 5 demonstrated that all of thecompounds of the present invention, represented by the test compound,showed a high inhibitory effect on FGFR1, FGFR3, and FGFR4, and servedas pan-FGFR inhibitors.

TABLE 23 IC₅₀ value (nM) Test compound Test Example 3 Test Example 4Test Example 5 Ex. Compound 2 3.6 0.5 3.4 Ex. Compound 5 1.4 0.2 0.5 Ex.Compound 9 3.2 0.3 4.9 Ex. Compound 47 20 0.5 11 Ex. Compound 51 1.2 0.11.7 Ex. Compound 52 2.8 0.7 21 Ex. Compound 62 3.1 0.7 19

Test Example 6

HEK293 cells were established in which a dovitinib-resistant FGFR2mutant N550H or E566G, or K660M, which is known as a mutation in uterinecancer, was expressed. Further, HEK293 cell lines were also establishedin which V565I, which is a mutation of the 565th valine at thegatekeeper site to isoleucine, was expressed. These cell lines wereincubated with the compound of Example 2 and further with a known FGFRinhibitor AZD4547 or BGJ-398. Then, the effect of inhibitingintracellular FGFR2 phosphorylation and cell growth was examined.

The results of Table 24 revealed that the inhibitory activity of bothAZD4547 and BGJ-398 on the dovitinib-resistant FGFR2 mutant N550H orE566G was less than that on wild-type FGFR2. The results demonstratedthat the mutants known to be resistant to the FGFR inhibitor dovitinibalso exhibited cross resistance to other FGFR inhibitors. It was shownthat the inhibitory activity of the compound of Example 2, which is anovel FGFR inhibitor, on these resistant FGFR2 mutants was equivalent tothat on the wild type. The inhibitory activity of both AZD4547 andBGJ-398 on the gatekeeper mutant V565I was significantly less than thaton the wild type; however, the compound of Example 2 maintainedinhibitory activity on this mutant almost equivalent to that on the wildtype. These results demonstrated that the compound of Example 2 hadinhibitory activity on the mutants resistant to dovitinib, AZD4547, andBGJ-398 equivalent to that on the wild type.

The same experiment was performed on K660M, which is a mutant found inuterine cancer. The results indicated that the inhibitory activity ofboth AZD4547 and BGJ-398 on this mutant was less than that on the wildtype, whereas the inhibitory activity of the compound of Example 2 onthis mutant was equivalent to that on the wild type. These resultsdemonstrated that the inhibitory activity of the compound of Example 2on tumor lines having FGFR mutations refractory to dovitinib, AZD4547,and BGJ-398 was equivalent to that on the wild-type FGFR2.

TABLE 24 pFGFR2 inhibition_IC₅₀ (nM) FGFR2 wild type N550H E566G K660MV565I Ex. 0.9 3.6 2.4 5.2 1.3 Compound 2 AZD4547 11.1 >300 189.1 167.093.8 BGJ398 6.0 >300 33.6 118.9 >3000

Test Example 7

Gastric cancer OCUM-2MD3 cells, for which FGFR2 gene amplification hadbeen reported, were cultured for about 3 months in the presence ofAZD4547, and cell clones resistant to this drug were isolated. FGFR2gene was isolated from the resistant clones, and the base sequence ofthe gene was decoded and compared with that of wild-type FGFR2. Further,the resistant cell lines were incubated with the compound of Example 2.Then, the effect of inhibiting intracellular FGFR2 phosphorylation andcell growth was examined.

Gastric cancer OCUM-2MD3 cells, for which FGFR2 gene amplification hadbeen reported, were cultured for about 3 months in the presence ofAZD4547, and a plurality of cell clones resistant to this drug wereisolated. When the base sequence of FGFR2 gene of these cell lines wasexamined, a mutation of the 660th amino acid K to N was observed. Thesecell lines were incubated with AZD4547, and the inhibition ofintracellular FGFR2 phosphorylation was examined. As a result, theinhibition in these cell lines was less than that in wild-type celllines. Mutations in the FGFR2 gene were considered to be one cause ofacquired resistance. When these clones were treated with the compound ofExample 2, FGFR2 phosphorylation and growth were inhibited to the sameextent as in the wild type (Tables 25 and 26).

TABLE 25 pFGFR2 inhibition, IC₅₀ (nM) Treatment Parent cell AZD4547resistant cells Ex. Compound 2 1.7  3.1 (range; 2.4-3.8) AZD4547 4.659.2 (33.0-94.3) BGJ398 4.5 22.6 (12.7-30.1)

TABLE 26 Cell growth inhibition, IC₅₀ (nM) Treatment Parent cell AZD4547resistant cells Ex. Compound 2 1.4  4.8 (range; 2.0-9.8) AZD4547 6.1158.6 (79.2-240.8) BGJ398 7.6  55.6 (33.9-98.7)Sequence Listing

The invention claimed is:
 1. A method of treating a patient having tumorcells resistant to an FGFR inhibitor, comprising administering aneffective amount of a 3,5-disubstituted benzene alkynyl compound or asalt thereof to the patient, the 3,5-disubstituted benzene alkynylcompound being represented by Formula (I):

wherein R₁ is the same or different, and each represents C₁-C₆ alkyl; X₁and X₂ independently represent N or CH; Y is a group represented byFormula (A):

wherein the divalent moiety represented by

is a nitrogen-containing C₃-C₁₀ heterocycloalkylene group, a grouprepresented by Formula (B):

wherein the divalent moiety represented by

is a C₃-C₁₀ cycloalkylene group, or a group represented by Formula (C):

wherein the divalent moiety represented by

is a C₆-C₁₂ arylene group; R₂ is hydrogen, C₂-C₆ alkynyl, —C(═O)OR_(x),—C(═O)N(R_(x))(R_(y)), hydroxy-C₁-C₆ alkyl, di(C₁-C₆ alkyl)amino-C₁-C₆alkyl, or C₂-C₉ heteroaryl optionally having R₃; and R₃ is C₁-C₆ alkylor di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl; Z is —C(R₄)═C(R₅)(R₆) or —C≡C—R₇;R₄, R₅, and R₆ are the same or different, and each represents hydrogen,halogen, C₁-C₆ alkyl optionally having R₈, or a group represented byFormula (D):

wherein the monovalent moiety represented by

is a nitrogen-containing C₃-C₁₀ heterocycloalkyl group, R₇ is hydrogen,C₁-C₆ alkyl, or hydroxy-C₁-C₆ alkyl; R₈ is —OR_(x) or —N(R_(x))(R_(y));R₉ is C₁-C₆ alkyl, halogen, or —OR_(x); R_(x) and R_(y) are the same ordifferent, and each represents hydrogen, C₁-C₆ alkyl, C₃-C₁₀ cycloalkyl,di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl, or C₁-C₆ alkoxy-C₁-C₆ alkyl; l is aninteger of 0 to 3; m is an integer of 1 to 3; and n is an integer of 0to 2, wherein the effective amount is 0.05 to 1,000 mg for an oralpreparation, 0.01 to 500 mg for an injection, and 1 to 1,000 mg for asuppository.
 2. The method according to claim 1, wherein the3,5-disubstituted benzene alkynyl compound is(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one.3. The method according to claim 1, wherein the patient has tumor cellsresistant to an FGFR inhibitor; and the tumor cells are resistant to atleast one of ponatinib, regorafenib, intedanib, dovitinib lactate,lenvatinib mesylate, cediranib, oratinib, brivanib alaninate, AZD4547,NVP-BGJ398, sulfatinib, ARQ-087, S-49076, IMCA1, PRO001, or R3Mab. 4.The method according to claim 1, wherein the patient has at least onemutation at a location selected from the group consisting of N550 ofFGFR2, V565 of FGFR2, E566 of FGFR2, and K660 of FGFR2.
 5. The methodaccording to claim 1, wherein the 3,5-disubstituted benzene alkynylcompound is administered orally.
 6. The method according to claim 1,wherein the patient is diagnosed with cancer selected from the groupconsisting of gastric cancer, liver cancer, cholangiocarcinoma, breastcancer, head and neck cancer, esophagus cancer, colon cancer, rectumcancer, gallbladder cancer, biliary tract cancer, pancreatic cancer,lung cancer, ovarian cancer, cervical cancer, endometrial cancer, renalcancer, bladder cancer, prostate cancer, testicular tumor, osteosarcoma,soft-tissue sarcoma, blood cancer, multiple myeloma, skin cancer, braintumor, and mesothelioma.
 7. The method according to claim 1, wherein thepatient is diagnosed with cholangiocarcinoma.
 8. The method according toclaim 1, wherein the 3,5-disubstituted benzene alkynyl compound is(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one;the patient has tumor cells resistant to at least one of AZD4547 orNVP-BGJ398; the cancer patient is diagnosed with gastric cancer; theeffective amount is 0.05 to 1,000 mg; and the 3,5-disubstituted benzenealkynyl compound is administered orally.
 9. A method of inhibiting FGFRkinase activity in a patient having tumor cells resistant to an FGFRinhibitor, comprising administering an effective amount of a3,5-disubstituted benzene alkynyl compound or a salt thereof to thepatient, the 3,5-disubstituted benzene alkynyl compound beingrepresented by Formula (I):

wherein R₁ is the same or different, and each represents C₁-C₆ alkyl; X₁and X₂ independently represent N or CH; Y is a group represented byFormula (A):

wherein the divalent moiety represented by

is a nitrogen-containing C₃-C₁₀ heterocycloalkylene group, a grouprepresented by Formula (B):

wherein the divalent moiety represented by

is a C₃-C₁₀ cycloalkylene group, or a group represented by Formula (C):

wherein the divalent moiety represented by

is a C₆-C₁₂ arylene group; R₂ is hydrogen, C₂-C₆ alkynyl, —C(═O)OR_(x),—C(═O)N(R_(x))(R_(y)), hydroxy-C₁-C₆ alkyl, di(C₁-C₆ alkyl)amino-C₁-C₆alkyl, or C₂-C₉ heteroaryl optionally having R₃; and R₃ is C₁-C₆ alkylor di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl; Z is —C(R₄)═C(R₅)(R₆) or —C≡C—R₇;R₄, R₅, and R₆ are the same or different, and each represents hydrogen,halogen, C₁-C₆ alkyl optionally having R₈, or a group represented byFormula (D):

wherein the monovalent moiety represented by

is a nitrogen-containing C₃-C₁₀ heterocycloalkyl group, R₇ is hydrogen,C₁-C₆ alkyl, or hydroxy-C₁-C₆ alkyl; R₈ is —OR_(x) or —N(R_(x))(R_(y));R₉ is C₁-C₆ alkyl, halogen, or —OR_(x); R_(x) and R_(y) are the same ordifferent, and each represents hydrogen, C₁-C₆ alkyl, C₃-C₁₀ cycloalkyl,di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl, or C₁-C₆ alkoxy-C₁-C₆ alkyl; l is aninteger of 0 to 3; m is an integer of 1 to 3; and n is an integer of 0to 2, wherein the effective amount is 0.05 to 1,000 mg for an oralpreparation, 0.01 to 500 mg for an injection, and 1 to 1,000 mg for asuppository.
 10. The method according to claim 9, wherein the3,5-disubstituted benzene alkynyl compound is(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one.11. The method according to claim 9, wherein the patient has tumor cellsthat are resistant to at least one of ponatinib, regorafenib, intedanib,dovitinib lactate, lenvatinib mesylate, cediranib, oratinib, brivanibalaninate, AZD4547, NVP-BGJ398, sulfatinib, ARQ-087, S-49076, IMCA1,PRO001, and R3Mab.
 12. The method according to claim 1, wherein thetumor cells resistant to the FGFR inhibitor are tumor cells thatresponded to a conventionally known FGFR inhibitor, but have becomeresistant to the antitumor effect of the inhibitor due to the continuousadministration thereof.
 13. The method according to claim 1, wherein thetumor cells resistant to the FGFR inhibitor are tumor cells that wereoriginally resistant to a conventionally known FGFR inhibitor.
 14. Themethod according to claim 9, wherein the tumor cells resistant to theFGFR inhibitor are tumor cells that responded to a conventionally knownFGFR inhibitor, but have become resistant to the antitumor effect of theinhibitor due to the continuous administration thereof.
 15. The methodaccording to claim 9, wherein the tumor cells resistant to the FGFRinhibitor are tumor cells that were originally resistant to aconventionally known FGFR inhibitor.
 16. A method of treating a patienthaving tumor cells resistant to an FGFR inhibitor, comprisingadministering an effective amount of a 3,5-disubstituted benzene alkynylcompound or a salt thereof to the patient, the 3,5-disubstituted benzenealkynyl compound being represented by Formula (I):

wherein R1 is the same or different, and each represents C₁-C₆ alkyl; X₁and X₂ are independently represent N or CH; Y is a group represented byFormula (A):

wherein the divalent moiety represented by

is a nitrogen-containing C₃-C₁₀ heterocycloalkylene group, a grouprepresented by Formula (B):

wherein the divalent moiety represented by

is a C₃-C₁₀ cycloalkylene group, or a group represented by Formula (C):

wherein the divalent moiety represented by

is a C₆-C₁₂ arylene group; R₂ is hydrogen, C₂-C₆ alkynyl, —C(═O)ORx,—C(═O)N(R_(x))(R_(y)), hydroxy-C₁-C₆ alkyl, di(C₁-C₆ alkyl)amino-C₁-C₆alkyl, or C₂-C₉ heteroaryl optionally having R₃; and R₃ is C₁-C₆ alkylor di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl; Z is —C(R₄)═C(R₅)(R₆) or —C═C—R₇;R₄, R₅, and R₆ are the same or different, and each represents hydrogen,halogen, C₁-C₆ alkyl optionally having R8, or a group represented byFormula (D):

wherein the monovalent moiety represented by

is a nitrogen-containing C₃-C₁₀ heterocycloalkyl group, R₇ is hydrogen,C₁-C₆ alkyl, or hydroxy-C₁-C₆ alkyl; R₈ is —OR_(x) or —N(R_(x))(R_(y));R₉ is C₁-C₆ alkyl, halogen, or —OR_(x); R_(x) and R_(y) are the same ordifferent, and each represents hydrogen, C₁-C₆ alkyl, C₃-C₁₀ cycloalkyl,di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl, or C₁-C₆ alkoxy-C₁-C₆ alkyl; l is aninteger of 0 to 3; m is an integer of 1 to 3; and n is an integer of 0to 2, wherein the patient is diagnosed with cancer selected from thegroup consisting of gastric cancer, liver cancer, cholangiocarcinoma,breast cancer, head and neck cancer, esophagus cancer, colon cancer,rectum cancer, gallbladder cancer, biliary tract cancer, pancreaticcancer, lung cancer, ovarian cancer, cervical cancer, endometrialcancer, renal cancer, bladder cancer, prostate cancer, testicular tumor,osteosarcoma, soft-tissue sarcoma, blood cancer, multiple myeloma, skincancer, brain tumor, and mesothelioma.
 17. The method according to claim16, wherein the 3,5-disubstituted benzene alkynyl compound is(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one.18. The method according to claim 16, wherein the patient has the tumorcells resistant to the FGFR inhibitor; and the tumor cells are resistantto at least one of ponatinib, regorafenib, intedanib, dovitinib lactate,lenvatinib mesylate, cediranib, oratinib, brivanib alaninate, AZD4547,NVP-BGJ398, sulfatinib, ARQ-087, S-49076, IMCA1, PRO001, and R3Mab. 19.The method according to claim 16, wherein the patient has at least onemutation at a location selected from the group consisting of N550 ofFGFR2, V565 of FGFR2, E566 of FGFR2, and K660 of FGFR2.
 20. The methodaccording to claim 16, wherein the 3,5-disubstituted benzene alkynylcompound is administered orally.
 21. The method according to claim 16,wherein the patient is diagnosed with cholangiocarcinoma.
 22. The methodaccording to claim 16, wherein the tumor cells resistant to the FGFRinhibitor are tumor cells that responded to a conventionally known FGFRinhibitor, but have become resistant to the antitumor effect of theinhibitor due to the continuous administration thereof.
 23. The methodaccording to claim 16, wherein the tumor cells resistant to the FGFRinhibitor are tumor cells that were originally resistant to aconventionally known FGFR inhibitor.
 24. The method according to claim16, wherein the 3,5-disubstituted benzene alkynyl compound is(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one;the patient has the tumor cells resistant to at least one of AZD4547 orNVP-BGJ398; the effective amount is 0.05 to 1,000 mg; the3,5-disubstituted benzene alkynyl compound is administered orally; andthe patient is diagnosed with gastric cancer.
 25. A method of inhibitingFGFR kinase activity in a patient having tumor cells resistant to anFGFR inhibitor, comprising administering an effective amount of a3,5-disubstituted benzene alkynyl compound or a salt thereof to thepatient, the 3,5-disubstituted benzene alkynyl compound beingrepresented by Formula (I):

wherein R₁ is the same or different, and each represents C₁-C₆ alkyl; X₁and X₂ independently represent N or CH; Y is a group represented byFormula (A):

wherein the divalent moiety represented by

is a nitrogen-containing C₃-C₁₀ heterocycloalkylene group, a grouprepresented by Formula (B):

wherein the divalent moiety represented by

is a C₃-C₁₀ cycloalkylene group, or a group represented by Formula (C):

wherein the divalent moiety represented by

is a C₆-C₁₂ arylene group; R₂ is hydrogen, C₂-C₆ alkynyl, —C(═O)OR_(x),—C(═O)N(R_(x))(R_(y)), hydroxy-C₁-C₆ alkyl, di(C₁-C₆ alkyl)amino-C₁-C₆alkyl, or C₂-C₉ heteroaryl optionally having R₃; and R₃ is C₁-C₆ alkylor di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl; Z is —C(R₄)═C(R₅)(R₆) or —C≡C—R₇;R₄, R₅, and R₆ are the same or different, and each represents hydrogen,halogen, C₁-C₆ alkyl optionally having R₈, or a group represented byFormula (D):

wherein the monovalent moiety represented by

is a nitrogen-containing C₃-C₁₀ heterocycloalkyl group, R₇ is hydrogen,C₁-C₆ alkyl, or hydroxy-C₁-C₆ alkyl; R₈ is —OR_(x) or —N(R_(x))(R_(y));R₉ is C₁-C₆ alkyl, halogen, or —OR_(x); Rx and Ry are the same ordifferent, and each represents hydrogen, C₁-C₆ alkyl, C₃-C₁₀ cycloalkyl,di(C₁-C₆ alkyl)amino-C₁-C₆ alkyl, or C₁-C₆ alkoxy-C₁-C₆ alkyl; l is aninteger of 0 to 3; m is an integer of 1 to 3; and n is an integer of 0to 2, wherein the patient is diagnosed with cancer selected from thegroup consisting of gastric cancer, liver cancer, cholangiocarcinoma,breast cancer, head and neck cancer, esophagus cancer, colon cancer,rectum cancer, gallbladder cancer, biliary tract cancer, pancreaticcancer, lung cancer, ovarian cancer, cervical cancer, endometrialcancer, renal cancer, bladder cancer, prostate cancer, testicular tumor,osteosarcoma, soft-tissue sarcoma, blood cancer, multiple myeloma, skincancer, brain tumor, and mesothelioma.
 26. The method according to claim25, wherein the 3,5-disubstituted benzene alkynyl compound is(S)-1-(3-(4-amino-3-((3,5-dimethoxyphenyl)ethynyl)-1H-pyrazolo[3,4-d]pyrimidin-1-yl)pyrrolidin-1-yl)prop-2-en-1-one.27. The method according to claim 25, wherein the patient has tumorcells that are resistant to at least one of ponatinib, regorafenib,intedanib, dovitinib lactate, lenvatinib mesylate, cediranib, oratinib,brivanib alaninate, AZD4547, NVP-BGJ398, sulfatinib, ARQ-087, S-49076,IMCA1, PRO001, or R3Mab.
 28. The method according to claim 25, whereinthe patient has at least one mutation at a location selected from thegroup consisting of N550 of FGFR2, V565 of FGFR2, E566 of FGFR2, andK660 of FGFR2.
 29. The method according to claim 25, wherein the tumorcells resistant to the FGFR inhibitor are tumor cells that responded toa conventionally known FGFR inhibitor, but have become resistant to theantitumor effect of the inhibitor due to the continuous administrationthereof.
 30. The method according to claim 25, wherein the tumor cellsresistant to the FGFR inhibitor are tumor cells that were originallyresistant to a conventionally known FGFR inhibitor.
 31. The methodaccording to claim 25, wherein the patient is diagnosed withcholangiocarcinoma.